Ergot Alkaloids of the Family Clavicipitaceae

PubMed Central

Florea, Simona; Panaccione, Daniel G.; Schardl, Christopher L.

2017-01-01

Ergot alkaloids are highly diverse in structure, exhibit diverse effects on animals, and are produced by diverse fungi in the phylum Ascomycota, including pathogens and mutualistic symbionts of plants. These mycotoxins are best known from the fungal family Clavicipitaceae and are named for the ergot fungi that, through millennia, have contaminated grains and caused mass poisonings, with effects ranging from dry gangrene to convulsions and death. However, they are also useful sources of pharmaceuticals for a variety of medical purposes. More than a half-century of research has brought us extensive knowledge of ergot-alkaloid biosynthetic pathways from common early steps to several taxon-specific branches. Furthermore, a recent flurry of genome sequencing has revealed the genomic processes underlying ergot-alkaloid diversification. In this review, we discuss the evolution of ergot-alkaloid biosynthesis genes and gene clusters, including roles of gene recruitment, duplication and neofunctionalization, as well as gene loss, in diversifying structures of clavines, lysergic acid amides, and complex ergopeptines. Also reviewed are prospects for manipulating ergot-alkaloid profiles to enhance suitability of endophytes for forage grasses. PMID:28168931

Transcriptome analysis of trichothecene-induced gene expression in barley.

PubMed

Boddu, Jayanand; Cho, Seungho; Muehlbauer, Gary J

2007-11-01

Fusarium head blight, caused primarily by Fusarium graminearum, is a major disease problem on barley (Hordeum vulgare L.). Trichothecene mycotoxins produced by the fungus during infection increase the aggressiveness of the fungus and promote infection in wheat (Triticum aestivum L.). Loss-of-function mutations in the TRI5 gene in F. graminearum result in the inability to synthesize trichothecenes and in reduced virulence on wheat. We examined the impact of pathogen-derived trichothecenes on virulence and the transcriptional differences in barley spikes infected with a trichothecene-producing wild-type strain and a loss-of-function tri5 trichothecene nonproducing mutant. Disease severity, fungal biomass, and floret necrosis and bleaching were reduced in spikes inoculated with the tri5 mutant strain compared with the wild-type strain, indicating that the inability to synthesize trichothecenes results in reduced virulence in barley. We detected 63 transcripts that were induced during trichothecene accumulation, including genes encoding putative trichothecene detoxification and transport proteins, ubiquitination-related proteins, programmed cell death-related proteins, transcription factors, and cytochrome P450s. We also detected 414 gene transcripts that were designated as basal defense response genes largely independent of trichothecene accumulation. Our results show that barley exhibits a specific response to trichothecene accumulation that can be separated from the basal defense response. We propose that barley responds to trichothecene accumulation by inducing at least two general responses. One response is the induction of genes encoding trichothecene detoxification and transport activities that may reduce the impact of trichothecenes. The other response is to induce genes encoding proteins associated with ubiquitination and cell death which may promote successful establishment of the disease.

Cytotoxicity and phytotoxicity of trichothecene mycotoxins produced by Fusarium spp.

PubMed

Abbas, Hamed K; Yoshizawa, Takumi; Shier, W Thomas

2013-11-01

Trichothecenes, a major class of mycotoxins produced by Fusarium, Myrothecium, and Stachybotrys species, are toxic to both plants and mammals. Simple trichothecenes, including type A (e.g., T-2 toxin) and type B (e.g., deoxynivalenol), are generally less toxic than macrocyclic trichothecenes. We sought to determine if simple trichothecenes are a potential source of candidates for development as bioherbicides, which require high phytotoxicity and low mammalian toxicity. We examined 28 simple trichothecenes in vitro for phytotoxicity using a small aquatic plant, Lemna pausicostata, and for mammalian toxicity using four cultured mammalian cell lines. Several structure-activity relationships were identified, including the following two, which may be relevant to bioherbicide development: peracetylation of type B trichothecenes and de-epoxidation of type A trichothecenes both substantially reduced mammalian toxicity with little effect on phytotoxicity. It was concluded that simple trichothecenes possessing strong phytotoxicity and minimal mammalian toxicity in vitro can be identified. Copyright © 2013 Elsevier Ltd. All rights reserved.

Ergot: from witchcraft to biotechnology.

PubMed

Haarmann, Thomas; Rolke, Yvonne; Giesbert, Sabine; Tudzynski, Paul

2009-07-01

The ergot diseases of grasses, caused by members of the genus Claviceps, have had a severe impact on human history and agriculture, causing devastating epidemics. However, ergot alkaloids, the toxic components of Claviceps sclerotia, have been used intensively (and misused) as pharmaceutical drugs, and efficient biotechnological processes have been developed for their in vitro production. Molecular genetics has provided detailed insight into the genetic basis of ergot alkaloid biosynthesis and opened up perspectives for the design of new alkaloids and the improvement of production strains; it has also revealed the refined infection strategy of this biotrophic pathogen, opening up the way for better control. Nevertheless, Claviceps remains an important pathogen worldwide, and a source for potential new drugs for central nervous system diseases.

Ergot and Its Alkaloids

PubMed Central

Schiff, Paul L.

2006-01-01

This manuscript reviews the history and pharmacognosy of ergot, and describes the isolation/preparation, chemistry, pharmacodynamics, and pharmacotherapeutics of the major ergot alkaloids and their derivatives. A brief discussion of the hallucinogenic properties of lysergic acid diethylamide is also featured. An abbreviated form of the material found in this paper is presented in a 4-hour didactic format to third-professional year PharmD students as part of their study of vascular migraine headaches, Parkinson's disease, and naturally occurring hallucinogens/hallucinogen derivatives in the modular course offering Neurology/Psychiatry. PMID:17149427

Analysis of Ergot Alkaloids

PubMed Central

Crews, Colin

2015-01-01

The principles and application of established and newer methods for the quantitative and semi-quantitative determination of ergot alkaloids in food, feed, plant materials and animal tissues are reviewed. The techniques of sampling, extraction, clean-up, detection, quantification and validation are described. The major procedures for ergot alkaloid analysis comprise liquid chromatography with tandem mass spectrometry (LC-MS/MS) and liquid chromatography with fluorescence detection (LC-FLD). Other methods based on immunoassays are under development and variations of these and minor techniques are available for specific purposes. PMID:26046699

QTL analysis of ergot resistance in sorghum.

PubMed

Parh, D K; Jordan, D R; Aitken, E A B; Mace, E S; Jun-ai, P; McIntyre, C L; Godwin, I D

2008-08-01

Sorghum ergot, caused predominantly by Claviceps africana Frederickson, Mantle, de Milliano, is a significant threat to the sorghum industry worldwide. The objectives of this study were firstly, to identify molecular markers linked to ergot resistance and to two pollen traits, pollen quantity (PQ) and pollen viability (PV), and secondly, to assess the relationship between the two pollen traits and ergot resistance in sorghum. A genetic linkage map of sorghum RIL population R931945-2-2 x IS 8525 (resistance source) was constructed using 303 markers including 36 SSR, 117 AFLP , 148 DArT and two morphological trait loci. Composite interval mapping identified nine, five, and four QTL linked to molecular markers for percentage ergot infection (PCERGOT), PQ and PV, respectively, at a LOD >2.0. Co-location/linkage of QTL were identified on four chromosomes while other QTL for the three traits mapped independently, indicating that both pollen and non pollen-based mechanisms of ergot resistance were operating in this sorghum population. Of the nine QTL identified for PCERGOT, five were identified using the overall data set while four were specific to the group data sets defined by temperature and humidity. QTL identified on SBI-02 and SBI-06 were further validated in additional populations. This is the first report of QTL associated with ergot resistance in sorghum. The markers reported herein could be used for marker-assisted selection for this important disease of sorghum.

Biosynthesis of trichothecenes and apotrichothecenes.

PubMed

Zamir, L O; Nikolakakis, A; Sauriol, F; Mamer, O

1999-05-01

Fusarium culmorum produces two major trichothecenes, 3-acetyldeoxynivalenol and sambucinol, and some minor apotrichothecenes. It was desired to investigate if during their biosynthesis a C-11-keto intermediate was involved. To verify this postulate, trichodiene, a known precursor to trichothecenes, was synthesized with two deuteriums at C-11 and one at C-15. It was then fed to F. culmorum cultures, and the derived metabolites were purified and analyzed. The results ruled out the involvement of an 11-keto intermediate but revealed two novel apotrichothecenes. The characterization of their structures suggested that one of the 2-hydroxy-11alpha-apotrichothecene stereoisomers (2alpha or 2beta) could be converted to sambucinol. These apotrichothecenes were therefore synthesized labeled specifically with two deuteriums at C-4 and C-15 and fed to F. culmorum cultures. Indeed, the result established for the first time that 2alpha-hydroxy-11alpha-apotrichothecene was a precursor to sambucinol. A biosynthetic scheme for the production of trichothecenes and apotrichothecenes is described.

Chemical and biological approaches for mycotoxin control: a review.

PubMed

Edlayne, Gonçalez; Simone, Aquino; Felicio, Joana D

2009-06-01

Mycotoxins are metabolites and toxic substances produced by certain filamentous fungi that frequently contaminate food and agriculture commodities, which cause disease in animals or man. The toxigenic fungi belong to mainly three genera: Aspergillus, Penicillium and Fusarium. Examples of mycotoxins of greatest public health and agroeconomic significance include aflatoxins, ochratoxins, trichothecenes, zearalenone, fumonisins, patulin and ergot alkaloids. Commodities susceptible to direct contamination with mycotoxins include nuts, oilseeds and grains. Chemical and biological treatments have been attempted to minimize the risk of mycotoxins contamination or eliminate the fungi of food and feeds. One way to prevent or interfere with fungal growth and mycotoxin production is by use of synthetic or natural agents. Bacteria have been studied to control the mycotoxins production and fungal growth in food. Plant genotypes resistant to infection by toxigenic fungi have been also studied. This review will approach same patented methods applied to degrade, prevent and control of mycotoxins in food and feeds.

Structure elucidation and in vitro cytotoxicity of ochratoxin α amide, a new degradation product of ochratoxin A.

PubMed

Bittner, Andrea; Cramer, Benedikt; Harrer, Henning; Humpf, Hans-Ulrich

2015-05-01

The mycotoxin ochratoxin A is a secondary metabolite occurring in a wide range of commodities. During the exposure of ochratoxin A to white and blue light, a cleavage between the carbon atom C-14 and the nitrogen atom was described. As a reaction product, the new compound ochratoxin α amide has been proposed based on mass spectrometry (MS) experiments. In the following study, we observed that this compound is also formed at high temperatures such as used for example during coffee roasting and therefore represents a further thermal ochratoxin A degradation product. To confirm the structure of ochratoxin α amide, the compound was prepared in large scale and complete structure elucidation via nuclear magnetic resonance (NMR) and MS was performed. Additionally, first studies on the toxicity of ochratoxin α amide were performed using immortalized human kidney epithelial (IHKE) cells, a cell line known to be sensitive against ochratoxin A with an IC50 value of 0.5 μM. Using this system, ochratoxin α amide revealed no cytotoxicity up to concentrations of 50 μM. Thus, these results propose that the thermal degradation of ochratoxin A to ochratoxin α amide might be a detoxification process. Finally, we present a sample preparation and a HPLC-tandem mass spectrometry (HPLC-MS/MS) method for the analysis of ochratoxin α amide in extrudates and checked its formation during the extrusion of artificially contaminated wheat grits at 150 and 180 °C, whereas no ochratoxin α amide was detectable under these conditions.

Cytotoxicity and accumulation of ergot alkaloids in human primary cells.

PubMed

Mulac, Dennis; Humpf, Hans-Ulrich

2011-04-11

Ergot alkaloids are secondary metabolites produced by fungi of the species Claviceps. Toxic effects after consumption of contaminated grains are described since mediaeval times. Of the more than 40 known ergot alkaloids six are found predominantly. These are ergotamine, ergocornine, ergocryptine, ergocristine, ergosine and ergometrine, along with their corresponding isomeric forms (-inine-forms). Toxic effects are known to be induced by an interaction of the ergot alkaloids as neurotransmitters, like dopamine or serotonin. Nevertheless data concerning cytotoxic effects are missing and therefore a screening of the six main ergot alkaloids was performed in human primary cells in order to evaluate the toxic potential. As it is well known that ergot alkaloids isomerize easily the stability was tested in the cell medium. Based on these results factors were calculated to correct the used concentration values to the biologically active lysergic (-ine) form. These factors range from 1.4 for the most stable compound ergometrine to 5.0 for the most unstable ergot alkaloid ergocristine. With these factors, reflecting the instability, several controverse literature data concerning the toxicity could be explained. To evaluate the cytotoxic effects of ergot alkaloids, human cells in primary culture were used. These cells remain unchanged in contrast to cell lines and the data allow a better comparison to the in vivo situation than using immortalized cell lines. To characterize the effects on primary cells, renal proximal tubule epithelial cells (RPTEC) and normal human astrocytes (NHA) were used. The parameters necrosis (LDH-release) and apoptosis (caspase-3-activation, DNA condensation and fragmentation) were distinguished. The results show that depending on the individual structure of the peptide ergot alkaloids the toxic properties change. While ergometrine as a lysergic acid amide did not show any effect, the peptide ergot alkaloids revealed a different toxic potential. Of

Protection against trichothecene mycotoxins. Final report

DOE Office of Scientific and Technical Information (OSTI.GOV)

Not Available

1983-01-01

Trichothecene mycotoxins, produced by certain fungi found in cereal grains, are a significant health problem in agricultural settings and have been detected throughout the world. Concern about the reported use of trichothecenes in chemical warfare agents, or 'yellow rain,' led the U.S. Army to request that the National Research Council form a committee on protection against mycotoxins, to study the effects of trichothecenes on civilians and military personnel who might be exposed to high levels of these substances. The report discusses scientific questions about the natural occurrences of mycotoxins, methods of detection and quantitation, decontamination and detoxification, long-term environmental effects,more » effects on humans and animals, and strategies for prevention and treatment. It also recommends areas of research that are particularly promising. An extensive bibliography is included.« less

Biosynthetic pathways of ergot alkaloids.

PubMed

Gerhards, Nina; Neubauer, Lisa; Tudzynski, Paul; Li, Shu-Ming

2014-12-10

Ergot alkaloids are nitrogen-containing natural products belonging to indole alkaloids. The best known producers are fungi of the phylum Ascomycota, e.g., Claviceps, Epichloë, Penicillium and Aspergillus species. According to their structures, ergot alkaloids can be divided into three groups: clavines, lysergic acid amides and peptides (ergopeptines). All of them share the first biosynthetic steps, which lead to the formation of the tetracyclic ergoline ring system (except the simplest, tricyclic compound: chanoclavine). Different modifications on the ergoline ring by specific enzymes result in an abundance of bioactive natural products, which are used as pharmaceutical drugs or precursors thereof. From the 1950s through to recent years, most of the biosynthetic pathways have been elucidated. Gene clusters from several ergot alkaloid producers have been identified by genome mining and the functions of many of those genes have been demonstrated by knock-out experiments or biochemical investigations of the overproduced enzymes.

An ergot alkaloid biosynthesis gene and clustered hypothetical genes from Aspergillus fumigatus.

PubMed

Coyle, Christine M; Panaccione, Daniel G

2005-06-01

The ergot alkaloids are a family of indole-derived mycotoxins with a variety of significant biological activities. Aspergillus fumigatus, a common airborne fungus and opportunistic human pathogen, and several fungi in the relatively distant taxon Clavicipitaceae (clavicipitaceous fungi) produce different sets of ergot alkaloids. The ergot alkaloids of these divergent fungi share a four-member ergoline ring but differ in the number, type, and position of the side chains. Several genes required for ergot alkaloid production are known in the clavicipitaceous fungi, and these genes are clustered in the genome of the ergot fungus Claviceps purpurea. We investigated whether the ergot alkaloids of A. fumigatus have a common biosynthetic and genetic origin with those of the clavicipitaceous fungi. A homolog of dmaW, the gene controlling the determinant step in the ergot alkaloid pathway of clavicipitaceous fungi, was identified in the A. fumigatus genome. Knockout of dmaW eliminated all known ergot alkaloids from A. fumigatus, and complementation of the mutation restored ergot alkaloid production. Clustered with dmaW in the A. fumigatus genome are sequences corresponding to five genes previously proposed to encode steps in the ergot alkaloid pathway of C. purpurea, as well as additional sequences whose deduced protein products are consistent with their involvement in the ergot alkaloid pathway. The corresponding genes have similarities in their nucleotide sequences, but the orientations and positions within the cluster of several of these genes differ. The data indicate that the ergot alkaloid biosynthetic capabilities in A. fumigatus and the clavicipitaceous fungi had a common origin.

An Ergot Alkaloid Biosynthesis Gene and Clustered Hypothetical Genes from Aspergillus fumigatus†

PubMed Central

Coyle, Christine M.; Panaccione, Daniel G.

2005-01-01

The ergot alkaloids are a family of indole-derived mycotoxins with a variety of significant biological activities. Aspergillus fumigatus, a common airborne fungus and opportunistic human pathogen, and several fungi in the relatively distant taxon Clavicipitaceae (clavicipitaceous fungi) produce different sets of ergot alkaloids. The ergot alkaloids of these divergent fungi share a four-member ergoline ring but differ in the number, type, and position of the side chains. Several genes required for ergot alkaloid production are known in the clavicipitaceous fungi, and these genes are clustered in the genome of the ergot fungus Claviceps purpurea. We investigated whether the ergot alkaloids of A. fumigatus have a common biosynthetic and genetic origin with those of the clavicipitaceous fungi. A homolog of dmaW, the gene controlling the determinant step in the ergot alkaloid pathway of clavicipitaceous fungi, was identified in the A. fumigatus genome. Knockout of dmaW eliminated all known ergot alkaloids from A. fumigatus, and complementation of the mutation restored ergot alkaloid production. Clustered with dmaW in the A. fumigatus genome are sequences corresponding to five genes previously proposed to encode steps in the ergot alkaloid pathway of C. purpurea, as well as additional sequences whose deduced protein products are consistent with their involvement in the ergot alkaloid pathway. The corresponding genes have similarities in their nucleotide sequences, but the orientations and positions within the cluster of several of these genes differ. The data indicate that the ergot alkaloid biosynthetic capabilities in A. fumigatus and the clavicipitaceous fungi had a common origin. PMID:15933009

Biosynthetic Pathways of Ergot Alkaloids

PubMed Central

Gerhards, Nina; Neubauer, Lisa; Tudzynski, Paul; Li, Shu-Ming

2014-01-01

Ergot alkaloids are nitrogen-containing natural products belonging to indole alkaloids. The best known producers are fungi of the phylum Ascomycota, e.g., Claviceps, Epichloë, Penicillium and Aspergillus species. According to their structures, ergot alkaloids can be divided into three groups: clavines, lysergic acid amides and peptides (ergopeptines). All of them share the first biosynthetic steps, which lead to the formation of the tetracyclic ergoline ring system (except the simplest, tricyclic compound: chanoclavine). Different modifications on the ergoline ring by specific enzymes result in an abundance of bioactive natural products, which are used as pharmaceutical drugs or precursors thereof. From the 1950s through to recent years, most of the biosynthetic pathways have been elucidated. Gene clusters from several ergot alkaloid producers have been identified by genome mining and the functions of many of those genes have been demonstrated by knock-out experiments or biochemical investigations of the overproduced enzymes. PMID:25513893

Genetics, Genomics and Evolution of Ergot Alkaloid Diversity

PubMed Central

Young, Carolyn A.; Schardl, Christopher L.; Panaccione, Daniel G.; Florea, Simona; Takach, Johanna E.; Charlton, Nikki D.; Moore, Neil; Webb, Jennifer S.; Jaromczyk, Jolanta

2015-01-01

The ergot alkaloid biosynthesis system has become an excellent model to study evolutionary diversification of specialized (secondary) metabolites. This is a very diverse class of alkaloids with various neurotropic activities, produced by fungi in several orders of the phylum Ascomycota, including plant pathogens and protective plant symbionts in the family Clavicipitaceae. Results of comparative genomics and phylogenomic analyses reveal multiple examples of three evolutionary processes that have generated ergot-alkaloid diversity: gene gains, gene losses, and gene sequence changes that have led to altered substrates or product specificities of the enzymes that they encode (neofunctionalization). The chromosome ends appear to be particularly effective engines for gene gains, losses and rearrangements, but not necessarily for neofunctionalization. Changes in gene expression could lead to accumulation of various pathway intermediates and affect levels of different ergot alkaloids. Genetic alterations associated with interspecific hybrids of Epichloë species suggest that such variation is also selectively favored. The huge structural diversity of ergot alkaloids probably represents adaptations to a wide variety of ecological situations by affecting the biological spectra and mechanisms of defense against herbivores, as evidenced by the diverse pharmacological effects of ergot alkaloids used in medicine. PMID:25875294

Feeding sorghum ergot (Claviceps africana) to sows before farrowing inhibits milk production.

PubMed

Kopinski, J S; Blaney, B J; Downing, J A; McVeigh, J F; Murray, S-A

2007-05-01

To assess the impact of feeding different amounts of sorghum ergot to sows before farrowing. Fifty-one pregnant sows from a continually farrowing piggery were sequentially inducted into the experiment each week in groups of four to seven, as they approached within 14 days of farrowing. Diets containing sorghum ergot sclerotia within the range of 0 (control) up to 1.5% w/w (1.5% ergot provided 7 mg alkaloids/kg, including 6 mg dihydroergosine/kg) were randomly allocated and individually fed to sows. Ergot concentrations were varied with each subsequent group until an acceptable level of tolerance was achieved. Diets with ergot were replaced with control diets after farrowing. Post-farrowing milk production was assessed by direct palpation and observation of udders, and by piglet responses and growth. Blood samples were taken from sows on three days each week, for prolactin estimation. Three sows fed 1.5% ergot for 6 to 10 days preceding farrowing produced no milk, and 87% of their piglets died despite supplementary feeding of natural and artificial colostrums, milk replacer, and attempts to foster them onto normally lactating sows. Ergot inclusions of 0.6% to 1.2% caused lesser problems in milk release and neo-natal piglet mortality. Of 23 sows fed either 0.3% or 0.6% ergot, lactation of only two first-litter sows were affected. Ergot caused pronounced reductions in blood prolactin, and first-litter sows had lower plasma prolactin than multiparous sows, increasing their susceptibility to ergot. Sorghum ergot should not exceed 0.3% (1 mg alkaloid/kg) in diets of multiparous sows fed before farrowing, and should be limited to 0.1% for primiparous sows, or avoided completely.

Antioxidant agents against trichothecenes: new hints for oxidative stress treatment

PubMed Central

Nepovimova, Eugenie; Wang, Yun; Yang, Hualin; Li, Li; Zhang, Xiujuan; Kuca, Kamil

2017-01-01

Trichothecenes are a group of mycotoxins mainly produced by fungi of genus Fusarium. Due to high toxicity and widespread dissemination, T-2 toxin and deoxynivalenol (DON) are considered to be the most important compounds of this class. Trichothecenes generate free radicals, including reactive oxygen species (ROS), which induce lipid peroxidation, decrease levels of antioxidant enzymes, and ultimately lead to apoptosis. Consequently, oxidative stress is an active area of research on the toxic mechanisms of trichothecenes, and identification of antioxidant agents that could be used against trichothecenes is crucial for human health. Numerous natural compounds have been analyzed and have shown to function very effectively as antioxidants against trichothecenes. In this review, we summarize the molecular mechanisms underlying oxidative stress induced by these compounds, and discuss current knowledge regarding such antioxidant agents as vitamins, quercetin, selenium, glucomannan, nucleotides, antimicrobial peptides, bacteria, polyunsaturated fatty acids, oligosaccharides, and plant extracts. These products inhibit trichothecene-induced oxidative stress by (1) inhibiting ROS generation and induced DNA damage and lipid peroxidation; (2) increasing antioxidant enzyme activity; (3) blocking the MAPK and NF-κB signaling pathways; (4) inhibiting caspase activity and apoptosis; (5) protecting mitochondria; and (6) regulating anti-inflammatory actions. Finally, we summarize some decontamination methods, including bacterial and yeast biotransformation and degradation, as well as mycotoxin-binding agents. This review provides a comprehensive overview of antioxidant agents against trichothecenes and casts new light on the attenuation of oxidative stress. PMID:29299181

On the Toxicity and Metabolism of the Trichothecene Mycotoxin T-2 Toxin,

DTIC Science & Technology

1988-06-20

on r~atiado d nfto..ayl --The present study deals witL1 toxicolegical effects anid :btoi of the trichothecene mycotoxin T-2 toxi.. T-2 toxin was shoeen...AND METABOLISM OF THE TRICHOTHECENE MYCOTOXIN T-2 TOXIN SUMMARY The present study deals with toxicological effects and metabolism of the trichothecene...directly related to this effect (McLaughlin et al., 1977). 15 1.4 DNA synthesis It is well established that trichothecene mycotoxins block DNA synthesis in

From yellow rain to green wheat: 25 years of trichothecene biosynthesis research.

PubMed

Desjardins, Anne E

2009-06-10

Trichothecene biosynthesis research at the U.S. Department of Agriculture in Peoria, IL, began in 1984 in response to concerns about the use of trichothecenes in biological warfare, but continued as a long-term research program on the intractable problem of trichothecene contamination of human foods and animal feeds. Over 25 years, the trichothecene biosynthesis research group integrated natural product chemistry with fungal genetics and plant pathology in the laboratory and in the field to understand how and why Fusarium species make these complex and highly toxic metabolites. This interdisciplinary research placed trichothecenes in the unique class of fungal metabolites that not only cause mycotoxicoses in animals but also are virulence factors in plant disease.

Ricinoleic acid as a marker for ergot impurities in rye and rye products.

PubMed

Franzmann, Carolin; Wächter, Johannes; Dittmer, Natascha; Humpf, Hans-Ulrich

2010-04-14

Ergot alkaloid and ricinoleic acid contents of 63 ergot sclerotia samples from rye throughout Germany of the harvest years 2006-2009 were determined. Alkaloid contents were analyzed by means of high-performance liquid chromatography with fluorescence detection (HPLC-FLD) and ricinoleic acid contents by means of gas chromatography with flame ionization detection (GC-FID). Ergot alkaloid amounts ranged from 0.01 to 0.2 g/100 g of sclerotia with an average amount of 0.08 g/100 g. Ergotamine and ergocristine were identified as lead alkaloids representing 57% (w/w) of the total alkaloid content. The average ricinoleic acid amount in the ergot sclerotia was 10.3 g/100 g. Because of the low variation of ricinoleic acid content in the ergot sclerotia, a new method for the determination of ricinoleic acid in rye products as a marker for ergot contaminations was developed. This method allows the determination of ergot impurities as low as 0.01% (w/w). Furthermore, 29 rye products (flours, bread mix, bread) were investigated for their ricinoleic acid and ergot alkaloid contents.

The history of ergot of rye (Claviceps purpurea) I: from antiquity to 1900.

PubMed

Lee, M R

2009-06-01

This article outlines the history of ergot of rye up to 1900. Ergot is a fungal disease that affects many grasses but is particularly damaging to rye. It occurs as the result of an infection by the parasitic organism Claviceps purpurea, which produces characteristic black spurs on the grass. When incorporated into grain, the ergot fungus can cause severe outbreaks of poisoning in humans called ergotism. There are two main clinical forms of toxicity, gangrenous and convulsive, where coma and death often supervene: the death rate for ergotism has been reported to be between 10 and 20 per cent in major outbreaks. Historical accounts note that ergot could accelerate labour, stop postpartum haemorrhage and inhibit lactation. At the end of the nineteenth century ergot was still regarded as a 'glorious chemical mess', but help would arrive in the early 1900s and the complex jigsaw would be solved.

Ergotism in Thailand caused by increased access to antiretroviral drugs: a global warning.

PubMed

Avihingsanon, Anchalee; Ramautarsing, Reshmie A; Suwanpimolkul, Gompol; Chetchotisakd, Ploenchan; Bowonwatanuwong, Chureeratana; Jirajariyavej, Supunnee; Kantipong, Patcharee; Tantipong, Hutsaya; Ohata, June Pirapon; Suankratay, Chusana; Ruxrungtham, Kiat; Burger, David M

2014-01-01

Ergotism is a toxic condition resulting from overexposure to the ergot compounds produced by various fungi of the genus Claviceps. Traditionally, such exposure was due to ingestion of infected grains, but long-term or excessive use of medications containing ergot derivatives or drug-drug interactions between these medications can result in ergotism. Ergotamine, typically used to treat migraine, has less than 5% bioavailability due to extensive first-pass metabolism by cytochrome P450 3A4 (CYP3A4). Concurrent intake of ergotamine and strong CYP3A4 inhibitors, such as the HIV protease inhibitors (PIs), can lead to clinical ergotism. A total of 13 cases of clinical ergotism in HIV-infected patients has been published since 1997 (most recently reviewed by Frohlich et al).

Effects of Trichothecenes on Cardiac Cell Electrical Function

DTIC Science & Technology

1985-12-16

toxic effects . These studies demonstrated unequivocal reversible effects of certain mycotoxins on heart cell electrical activity. Preliminary studies...muscle cells shown in Figure 8 illustrate the typical effects of trichothecene mycotoxins in canine ventricular cells. T-2 tetraol, for 3xample...false tendon cells and V ventricular muscle cells (shown in Figure 8) illustrate the typical effects of trichothecene mycotoxins in canine cardiac

Rapid Screening of Ergot Alkaloids in Sclerotia by MALDI-TOF Mass Spectrometry.

PubMed

Sivagnanam, Kumaran; Komatsu, Emy; Patrick, Susan; Rampitsch, Christoph; Perreault, Hélène; Gräfenhan, Tom

2016-07-01

Ergot is a common disease of wheat and other cereal grains that is predominantly caused by Claviceps purpurea in the field, often affecting crop yield in addition to the environment. Infected grain can be contaminated with dark sclerotia, which contain fungal metabolites such as ergot alkaloids. The occurrence of ergot alkaloids in cereal grain is a major health concern for humans and livestock. Effective and rapid screening of these mycotoxins is crucial for producers, processors, and consumers of cereal-based food and feed grain. Established methods of ergot alkaloid screening based on LC-MS or GC-MS require laborious processes. A novel method using matrix-assisted laser desorption ionization (MALDI)-time-of-flight (TOF) MS was developed to identify four ergot alkaloids. Using dihydroxybenzoic acid as the matrix, ergosine, ergocornine, ergocryptine, and ergocristine were readily detected in individual sclerotia of C. purpurea. The accuracy of the identified ergot alkaloids was further confirmed by tandem MS analysis. MALDI-TOF MS is suitable for high-throughput screening of ergot alkaloids because it permits rapid and accurate identification, simple sample preparation, and no derivatization or chromatographic separation.

Is the pathogenic ergot fungus a conditional defensive mutualist for its host grass?

PubMed

Wäli, Pauliina P; Wäli, Piippa R; Saikkonen, Kari; Tuomi, Juha

2013-01-01

It is well recognized, that outcomes of mutualistic plant-microorganism interactions are often context dependent and can range from mutualistic to antagonistic depending on conditions. Instead, seemingly pathogenic associations are generally considered only harmful to plants. The ergot fungus (Claviceps purpurea) is a common seed pathogen of grasses and cereals. Ergot sclerotia contain alkaloids which can cause severe toxicity in mammals when ingested, and thus the fungal infection might provide protection for the host plant against mammalian herbivores. Theoretically, the net effect of ergot infection would positively affect host seed set if the cost is not too high and the defensive effect is strong enough. According to our empirical data, this situation is plausible. First, we found no statistically significant seed loss in wild red fescue (Festuca rubra) inflorescences due to ergot infection, but the seed succession decreased along increasing number of sclerotia. Second, in a food choice experiment, sheep showed avoidance against forage containing ergot. Third, the frequency of ergot-infected inflorescences was higher in sheep pastures than surrounding ungrazed areas, indicating a protective effect against mammalian grazing. We conclude that, although ergot can primarily be categorized as a plant pathogen, ergot infection may sometimes represent indirect beneficial effects for the host plant. Ergot may thus serve as a conditional defensive mutualist for its host grass, and the pathogenic interaction may range from antagonistic to mutualistic depending on the situation.

Effects of two metabolites of ochratoxin A, (4R)-4-hydroxyochratoxin A and ochratoxin alpha, on immune response in mice.

PubMed Central

Creppy, E E; Størmer, F C; Röschenthaler, R; Dirheimer, G

1983-01-01

The metabolites of ochratoxin A, (4R)-4-hydroxyochratoxin A and ochratoxin alpha, were investigated for immunosuppressive properties in BALB/c mice. The standard plaque-counting technique for the estimation of antibody-producing spleen lymphocytes was used. (4R)-4-hydroxyochratoxin A was found to be an immunosuppressor almost as highly effective as ochratoxin A. Doses of 1 microgram of (4R)-4-hydroxyochratoxin A per kg administered to mice caused an 80% reduction in the number of cells producing immunoglobulin M (90% with ochratoxin A) and a 93% reduction in cells synthesizing immunoglobulin G (92% with ochratoxin A). Ochratoxin alpha, however, was ineffective. A possible mode of action is discussed. PMID:6341224

Post-genome research on the biosynthesis of ergot alkaloids.

PubMed

Li, Shu-Ming; Unsöld, Inge A

2006-10-01

Genome sequencing provides new opportunities and challenges for identifying genes for the biosynthesis of secondary metabolites. A putative biosynthetic gene cluster of fumigaclavine C, an ergot alkaloid of the clavine type, was identified in the genome sequence of ASPERGILLUS FUMIGATUS by a bioinformatic approach. This cluster spans 22 kb of genomic DNA and comprises at least 11 open reading frames (ORFs). Seven of them are orthologous to genes from the biosynthetic gene cluster of ergot alkaloids in CLAVICEPS PURPUREA. Experimental evidence of the identified cluster was provided by heterologous expression and biochemical characterization of two ORFs, FgaPT1 and FgaPT2, in the cluster of A. FUMIGATUS, which show remarkable similarities to dimethylallyltryptophan synthase from C. PURPUREA and function as prenyltransferases. FgaPT2 converts L-tryptophan to dimethylallyltryptophan and thereby catalyzes the first step of ergot alkaloid biosynthesis, whilst FgaPT1 catalyzes the last step of the fumigaclavine C biosynthesis, i. e., the prenylation of fumigaclavine A at C-2 position of the indole nucleus. In addition to information obtained from the gene cluster of ergot alkaloids from C. PURPUREA, the identification of the biosynthetic gene cluster of fumigaclavine C in A. FUMIGATUS opens an alternative way to study the biosynthesis of ergot alkaloids in fungi.

Phytotoxic effects of trichothecenes on the growth and morphology of Arabidopsis thaliana.

PubMed

Masuda, Daisuke; Ishida, Mamoru; Yamaguchi, Kazuo; Yamaguchi, Isamu; Kimura, Makoto; Nishiuchi, Takumi

2007-01-01

Non-volatile sesquiterpenoids, a trichothecene family of phytotoxins such as deoxynivalenol (DON) and T-2 toxin, contain numerous molecular species and are synthesized by phytopathogenic Fusarium species. Although trichothecene chemotypes might play a role in the virulence of individual Fusarium strains, the phytotoxic action of individual trichothecenes has not been systematically studied. To perform a comparative analysis of the phytotoxic action of representative trichothecenes, the growth and morphology of Arabidopsis thaliana growing on media containing these compounds was investigated. Both DON and diacetoxyscirpenol (DAS) preferentially inhibited root elongation. DON-treated roots were less organized compared with control roots. Moreover, preferential inhibition of root growth by DON was also observed in wheat plants. In addition, T-2 toxin-treated seedlings exhibited dwarfism with aberrant morphological changes (e.g. petiole shortening, curled dark-green leaves, and reduced cell size). These results imply that the phytotoxic action of trichothecenes differed among their molecular species. Cycloheximide (CHX)-treated seedlings displayed neither feature, although it is known that trichothecenes inhibit translation in eukaryotic ribosomes. Microarray analyses suggested that T-2 toxin caused a defence response, the inactivation of brassinosteroid (BR), and the generation of reactive oxygen species in Arabidopsis. This observation is in agreement with our previous reports in which trichothecenes such as T-2 toxin have an elicitor-like activity when infiltrated into the leaves of Arabidopsis. Since it has been reported that BR plays an important role in a broad range of disease resistance in tobacco and rice, inactivation of BR might affect pathogenicity during the infection of host plants by trichothecene-producing fungi.

Is the Pathogenic Ergot Fungus a Conditional Defensive Mutualist for Its Host Grass?

PubMed Central

Wäli, Pauliina P.; Wäli, Piippa R.; Saikkonen, Kari; Tuomi, Juha

2013-01-01

It is well recognized, that outcomes of mutualistic plant-microorganism interactions are often context dependent and can range from mutualistic to antagonistic depending on conditions. Instead, seemingly pathogenic associations are generally considered only harmful to plants. The ergot fungus (Claviceps purpurea) is a common seed pathogen of grasses and cereals. Ergot sclerotia contain alkaloids which can cause severe toxicity in mammals when ingested, and thus the fungal infection might provide protection for the host plant against mammalian herbivores. Theoretically, the net effect of ergot infection would positively affect host seed set if the cost is not too high and the defensive effect is strong enough. According to our empirical data, this situation is plausible. First, we found no statistically significant seed loss in wild red fescue (Festuca rubra) inflorescences due to ergot infection, but the seed succession decreased along increasing number of sclerotia. Second, in a food choice experiment, sheep showed avoidance against forage containing ergot. Third, the frequency of ergot-infected inflorescences was higher in sheep pastures than surrounding ungrazed areas, indicating a protective effect against mammalian grazing. We conclude that, although ergot can primarily be categorized as a plant pathogen, ergot infection may sometimes represent indirect beneficial effects for the host plant. Ergot may thus serve as a conditional defensive mutualist for its host grass, and the pathogenic interaction may range from antagonistic to mutualistic depending on the situation. PMID:23874924

Genome mining of ascomycetous fungi reveals their genetic potential for ergot alkaloid production.

PubMed

Gerhards, Nina; Matuschek, Marco; Wallwey, Christiane; Li, Shu-Ming

2015-06-01

Ergot alkaloids are important as mycotoxins or as drugs. Naturally occurring ergot alkaloids as well as their semisynthetic derivatives have been used as pharmaceuticals in modern medicine for decades. We identified 196 putative ergot alkaloid biosynthetic genes belonging to at least 31 putative gene clusters in 31 fungal species by genome mining of the 360 available genome sequences of ascomycetous fungi with known proteins. Detailed analysis showed that these fungi belong to the families Aspergillaceae, Clavicipitaceae, Arthrodermataceae, Helotiaceae and Thermoascaceae. Within the identified families, only a small number of taxa are represented. Literature search revealed a large diversity of ergot alkaloid structures in different fungi of the phylum Ascomycota. However, ergot alkaloid accumulation was only observed in 15 of the sequenced species. Therefore, this study provides genetic basis for further study on ergot alkaloid production in the sequenced strains.

Analysis and modification of ergot alkaloid profiles in fungi.

PubMed

Panaccione, Daniel G; Ryan, Katy L; Schardl, Christopher L; Florea, Simona

2012-01-01

The ergot alkaloids are a family of secondary metabolites produced by a phylogenetically discontinuous group of fungi. Various members of the family are important in agriculture, where they accumulate in grain crops or forage grasses and adversely affect humans or animals who consume them. Other ergot alkaloids have been used clinically to treat a variety of diseases. Because of their significance in agriculture and medicine, the ability to detect and quantify these alkaloids from a variety of substrates is important. The primary analytical approach for these purposes has been high performance liquid chromatography. The ability to manipulate ergot alkaloid production in fungi, by transformation-mediated approaches, has been useful for studies on the biosynthesis of these alkaloids and may have practical application in agriculture and medicine. Such modifications have been informed by comparative genomic approaches, which have provided information on the gene clusters associated with ergot alkaloid biosynthesis. Copyright © 2012 Elsevier Inc. All rights reserved.

Chronic ergot toxicity presenting with bilateral external iliac artery dissection and lower extremity rest pain.

PubMed

Molkara, Afshin M; Abou-Zamzam, Ahmed M; Teruya, Theodore H; Bianchi, Christian; Killeen, J David

2006-11-01

Chronic use of ergot alkaloids has been recognized as a rare cause of lower extremity ischemia. Most patients with ergot toxicity present with symptoms of lower extremity claudication. Herein we present a woman with bilateral lower extremity rest pain and a history of chronic ergot use for migraine headaches. Arteriography demonstrated extensive pruning of the distal arterial tree along with bilateral external iliac artery dissections - a finding that is not often associated with young, normotensive patients with chronic ergot toxicity. This patient was treated with endovascular stenting of the dissections along with cessation of ergot. Her symptoms improved markedly, and follow-up arteriography 6 weeks later demonstrated resolution of the iliac dissections along with restoration of nearly normal lower extremity runoff vessels. Discontinuation of ergot-containing products and cessation of tobacco and caffeine use is the cornerstone of therapy in chronic ergot toxicity. The association of ergot toxicity and iliac dissection has not been previously described. Endovascular or surgical interventions may be considered in patients with ergot toxicity for specific indications or those whose symptoms progress despite conservative management.

Analysis of Trichothecene Mycotoxins by Combined HPLC/MS.

DTIC Science & Technology

1986-04-15

trichothecenes are known and several of them are natural contaminants of cereal grains(l). Ingestion of the contaminated foodstuffs by humans or farm...71.2% of the total when urine, which contains naturally occurring salts, was used instead of distilled water. The use of saturated sodium chloride...Imperfecti. Many trichothecenes are known and several of them are natural contaminants of cereal grains(l). Ingestion of the contaminated foodstuffs by

Hypothesis: does ochratoxin A cause testicular cancer?

PubMed

Schwartz, Gary G

2002-02-01

Little is known about the etiology of testicular cancer, which is the most common cancer among young men. Epidemiologic data point to a carcinogenic exposure in early life or in utero, but the nature of the exposure is unknown. We hypothesize that the mycotoxin, ochratoxin A, is a cause of testicular cancer. Ochratoxin A is a naturally occurring contaminant of cereals, pigmeat, and other foods and is a known genotoxic carcinogen in animals. The major features of the descriptive epidemiology of testicular cancer (a high incidence in northern Europe, increasing incidence over time, and associations with high socioeconomic status, and with poor semen quality) are all associated with exposure to ochratoxin A. Exposure of animals to ochratoxin A via the diet or via in utero transfer induces adducts in testicular DNA. We hypothesize that consumption of foods contaminated with ochratoxin A during pregnancy and/or childhood induces lesions in testicular DNA and that puberty promotes these lesions to testicular cancer. We tested the ochratoxin A hypothesis using ecologic data on the per-capita consumption of cereals, coffee, and pigmeat, the principal dietary sources of ochratoxin A. Incidence rates for testicular cancer in 20 countries were significantly correlated with the per-capita consumption of coffee and pigmeat (r = 0.49 and 0.54, p = 0.03 and 0.01). The ochratoxin A hypothesis offers a coherent explanation for much of the descriptive epidemiology of testicular cancer and suggests new avenues for analytic research.

[Ergotismus convulsivus. the convulsive type of ergotism illustrated by an example in the year 1738].

PubMed

Priewer, Helmut; Priewer, Mathias

2010-01-01

This paper describes cases of ergotismus convulsivus (the kind of poisoning from ergotized grain marked by convulsions), some of them fatal, in the year 1738. The origins of the formation of the ergotized grain as well as the symptoms of ergotism are presented. Comparisons to other epidemics of ergotism are drawn.

Effects of Trichothecenes on Cardiac Cell Electrical Function

DTIC Science & Technology

1985-12-13

Figure 8 illustrate the typical effects of trichothecene mycotoxins in canine ventricular cells. T-2 tetraol, for example, reduced the total duration of...potentials from false tendon cells and ventricular muscle cells (shown in Figure 8) illustrate the typical effects of trichothecene mycotoxins in canine...the plateau (arrow) from 14 my to 4 my. Table 6 summarizes the effects of T-2 mycotoxin on the action potential parameters of false tendon cells and

Tall fescue seed extraction and partial purification of ergot alkaloids

USDA-ARS?s Scientific Manuscript database

Many substances in the tall fescue/endophyte association (Schedonorus arundinaceus/Epichloë coenophiala) have biological activity. Of these compounds only the ergot alkaloids are known to have significant mammalian toxicity and the predominant ergot alkaloids are ergovaline and ergovalinine. Because...

Ergot Alkaloids and their Hallucinogenic Potential in Morning Glories.

PubMed

Steiner, Ulrike; Leistner, Eckhard

2018-03-02

Naturally occurring and semisynthetic ergot alkaloids play a role in health care or as recreational drugs in Western and indigenous Mexican societies. Evidence is summarized that ergot alkaloids present in Central American Convolvulaceae like Turbina corymbosa, Ipomoea violacea , and Ipomoea asarifolia are colonized by different species of a newly described clavicipitaceous fungal genus named Periglandula . The fungi are associated with peltate glandular trichomes on the adaxial leaf surface of its host plants. The Periglandula fungi are not yet culturable in vitro but were demonstrated to have the capacity to synthesize ergot alkaloids. The alkaloids do not remain in the fungal mycelium but are translocated via the glandular trichomes into their plant host. Both fungi and host benefit from a symbiotic lifestyle. In evolutionary terms the alkaloid biosynthetic gene cluster in the Periglandula/Ipomoea symbiosis is likely to have a conserved (basic) structure while biosynthetic ergot gene clusters within the genera Claviceps and Epichloe were under ecological selection for alkaloid diversification. Georg Thieme Verlag KG Stuttgart · New York.

Endophyte-associated ergot alkaloids

USDA-ARS?s Scientific Manuscript database

Fescue toxicosis is a very costly (greater than $600 million/annually) for the cattle, horse and small ruminant industries. The tall fescue forage responsible for this intoxication is infected with an endophytic fungus (Neotyphodium coenophialum) that produces ergot alkaloids, which are toxic to th...

Ergot Alkaloids in Feed for Pekin Ducks: Toxic Effects, Metabolism and Carry Over into Edible Tissues

PubMed Central

Dänicke, Sven

2015-01-01

Hardened sclerotia (ergots) of Claviceps purpurea contaminate cereal grains and contain toxic ergot alkaloids (EA). Information on EA toxicity in ducks is scarce. Therefore, the aim of the growth experiment (Day 0–49, n = 54/group) was to titrate the lowest observed adverse effect level (LOAEL) for total ergot alkaloids (TEA). A control diet was prepared without ergots, and the diets designated Ergot 1 to 4 contained 1, 10, 15 and 20 g ergot per kg diet, respectively, corresponding to TEA contents of 0.0, 0.6, 7.0, 11.4 and 16.4 mg/kg. Sensitivity of ducks to EA was most pronounced at the beginning of the experiment when feed intake decreased significantly by 9%, 28%, 41% and 47% in groups Ergot 1 to 4, respectively, compared to the control group. The experiment was terminated after two weeks for ducks exposed to Ergot 3 and 4 due to significant growth retardation. Ergot alkaloid residues in edible tissues were lower than 5 ng/g. Bile was tested positive for ergonovine (=ergometrine = ergobasine) with a mean concentration of 40 ng/g. Overall, the LOAEL amounted to 0.6 mg TA/kg diet suggesting that ducks are not protected by current European Union legislation (1 g ergot/kg unground cereal grains). PMID:26043275

Ergot alkaloids in feed for Pekin ducks: toxic effects, metabolism and carry over into edible tissues.

PubMed

Dänicke, Sven

2015-06-02

Hardened sclerotia (ergots) of Claviceps purpurea contaminate cereal grains and contain toxic ergot alkaloids (EA). Information on EA toxicity in ducks is scarce. Therefore, the aim of the growth experiment (Day 0-49, n = 54/group) was to titrate the lowest observed adverse effect level (LOAEL) for total ergot alkaloids (TEA). A control diet was prepared without ergots, and the diets designated Ergot 1 to 4 contained 1, 10, 15 and 20 g ergot per kg diet, respectively, corresponding to TEA contents of 0.0, 0.6, 7.0, 11.4 and 16.4 mg/kg. Sensitivity of ducks to EA was most pronounced at the beginning of the experiment when feed intake decreased significantly by 9%, 28%, 41% and 47% in groups Ergot 1 to 4, respectively, compared to the control group. The experiment was terminated after two weeks for ducks exposed to Ergot 3 and 4 due to significant growth retardation. Ergot alkaloid residues in edible tissues were lower than 5 ng/g. Bile was tested positive for ergonovine (=ergometrine = ergobasine) with a mean concentration of 40 ng/g. Overall, the LOAEL amounted to 0.6 mg TA/kg diet suggesting that ducks are not protected by current European Union legislation (1 g ergot/kg unground cereal grains).

Toxigenic Fusarium spp. as determinants of trichothecene mycotoxins in settled grain dust.

PubMed

Halstensen, Anne Straumfors; Nordby, Karl-Christian; Klemsdal, Sonja Sletner; Elen, Oleif; Clasen, Per-Erik; Eduard, Wijnand

2006-12-01

Trichothecenes are immunosuppressive mycotoxins produced mainly by Fusarium spp. and often are detected as natural contaminants of grain and other agricultural products. Exposure to trichothecenes through inhalation during grain work may represent possible health risks for grain farmers. We aimed, therefore, to investigate the level of Fusarium spp. and trichothecenes in settled grain dust collected during work on 92 Norwegian farms. Mycotoxins were determined by gas chromatography-mass spectrometry, whereas the Fusarium spp. were identified and quantified both by species-specific semiquantitative polymerase chain reaction (PCR) and by cultivation. All potential trichothecene-producing molds in the grain dust were quantified using a PCR assay specific for tri5, the gene coding for trichodiene synthase that catalyzes the first step in the trichothecene biosynthesis. We performed correlation analysis between mold-DNA and mycotoxins to assess whether the PCR-detected DNA could be used as indicators of the mycotoxins. The methodological problem of detecting small amounts of airborne mycotoxins during grain work may then be avoided. Whereas the trichothecene-producing Fusarium species in grain dust could not be identified or quantified to a sufficient extent by cultivation, all investigated Fusarium spp. could be specifically detected by PCR and quantified from the DNA agarose gel band intensities. Furthermore, we observed a strong correlation between the trichothecenes HT-2 toxin (HT-2) or T-2 toxin (T-2) and DNA specific for tri5 (r = 0.68 for HT-2 and r = 0.50 for T-2; p < 0.001), F. langsethiae (r = 0.77 for HT-2 and r = 0.59 for T-2; p < 0.001), or F. poae (r = 0.41 for HT-2 and r = 0.35 for T-2; p < 0.001). However, only a moderate correlation was observed between the trichothecene deoxynivalenol (DON) and the combination of its producers, F. culmorum and F. graminearum (r = 0.24, p = 0.02), and no significant correlation was observed between DON and tri5. PCR

Effect of feeding fescue seed containing ergot alkaloid toxins on stallion spermatogenesis and sperm cells.

PubMed

Fayrer-Hosken, R; Stanley, A; Hill, N; Heusner, G; Christian, M; De La Fuente, R; Baumann, C; Jones, L

2012-12-01

The cellular effects of tall fescue grass-associated toxic ergot alkaloids on stallion sperm and colt testicular tissue were evaluated. This was a continuation of an initial experiment where the effects of toxic ergot alkaloids on the stallion spermiogram were investigated. The only spermiogram parameter in exposed stallions that was affected by the toxic ergot alkaloids was a decreased gel-free volume of the ejaculate. This study examined the effect of toxic ergot alkaloids on chilling and freezing of the stallion sperm cells. The effect of toxic ergot alkaloids on chilled extended sperm cells for 48 h at 5°C was to make the sperm cells less likely to undergo a calcium ionophore-induced acrosome reaction. The toxic ergot alkaloids had no effect on the freezability of sperm cells. However, if yearling colts were fed toxic ergot alkaloids, then the cytological analysis of meiotic chromosome synapsis revealed a significant increase in the proportion of pachytene spermatocytes showing unpaired sex chromosomes compared to control spermatocytes. There was little effect of ergot alkaloids on adult stallions, but there might be a significant effect on yearling colts. © 2012 Blackwell Verlag GmbH.

Effect of Feeding Fescue Seed Containing Ergot Alkaloid Toxins on Stallion Spermatogenesis and Sperm Cells

PubMed Central

Fayrer-Hosken, R; Stanley, A; Hill, N; Heusner, G; Christian, M; Fuente, R De La; Baumann, C; Jones, L

2012-01-01

Contents The cellular effects of tall fescue grass–associated toxic ergot alkaloids on stallion sperm and colt testicular tissue were evaluated. This was a continuation of an initial experiment where the effects of toxic ergot alkaloids on the stallion spermiogram were investigated. The only spermiogram parameter in exposed stallions that was affected by the toxic ergot alkaloids was a decreased gel-free volume of the ejaculate. This study examined the effect of toxic ergot alkaloids on chilling and freezing of the stallion sperm cells. The effect of toxic ergot alkaloids on chilled extended sperm cells for 48 h at 5 °C was to make the sperm cells less likely to undergo a calcium ionophore–induced acrosome reaction. The toxic ergot alkaloids had no effect on the freezability of sperm cells. However, if yearling colts were fed toxic ergot alkaloids, then the cytological analysis of meiotic chromosome synapsis revealed a significant increase in the proportion of pachytene spermatocytes showing unpaired sex chromosomes compared to control spermatocytes. There was little effect of ergot alkaloids on adult stallions, but there might be a significant effect on yearling colts. PMID:22524585

Production of trichothecene mycotoxins by Australian Fusarium species.

PubMed

McLachlan, A; Shaw, K J; Hocking, A D; Pitt, J I; Nguyen, T H

1992-01-01

Australian isolates of Fusarium species were grown on potato dextrose agar. Trichothecenes produced by these species were extracted by ethyl acetate followed by methanol and a silica gel column was used to clean-up the extract. The extracted samples were derivatized by acetylation with trifluoroacetic anhydride and the derivatives analysed by gas chromatography/mass spectrometry (GC/MS). Multiple ion detection was used to trace ions characteristic of the trichothecenes expected to be present. Quantitation of those found was based on a known mass of pentabromophenol that was added as an internal standard. Eight species of Fusarium (nineteen strains) were surveyed, of which three species, F. acuminatum, F. equiseti and F. sporotrichioides, produced the trichothecenes scirpentriol, diacetoxyscirpenol, neosolaniol, HT-2 toxin, T-2 toxin, T-2 tetraol and deoxynivalenol. Wheat samples were inoculated with four different species of Fusarium, F. acuminatum, F. equiseti, F. graminearum and F. sporotrichioides, and in these samples diacetoxyscirpenol, neosolaniol, HT-2 toxin and T-2 toxin were found.

Mycotoxin occurrence in commodities, feeds and feed ingredients sourced in the Middle East and Africa

PubMed Central

Rodrigues, I.; Handl, J.; Binder, E.M.

2011-01-01

Between February and October 2009, 324 grain, feed and feed commodity samples were sourced directly at animal farms or feed production sites in Middle East and Africa and tested for the presence of A- and B-trichothecenes, zearalenone, fumonisins, aflatoxins and ochratoxin A, or for selected groups of mycotoxins only. Samples were analyzed after clean-up by immunoaffinity or solid-phase extraction followed by HPLC with derivatization where appropriate and fluorescence, UV or mass spectrometric detection. The percentage of positive samples of B-trichothecenes ranged from 0 to 87% of tested samples. The prevalence of fumonisins in the different countries was >50% in most cases. Zearalenone was present in tested commodities from all countries except three. The presence of aflatoxin in analyzed samples varied from 0 to 94%. Ochratoxin A was present in 67% of samples in Sudan and in 100% of Nigerian samples. No A-trichothecenes were found in this survey. PMID:24786003

Biodegradation of ochratoxin A by Alcaligenes faecalis isolated from soil.

PubMed

Zhang, H H; Wang, Y; Zhao, C; Wang, J; Zhang, X L

2017-09-01

The aim of this study is to report on the hydrolytic action of Alcaligenes faecalis isolated from soil samples and its ability to degrade ochratoxin A. An A. faecalis strain was identified and characterized by employing both a phenotypic analysis and 16S rDNA sequence analysis. The results show that this strain could degrade ochratoxin A efficiently but could not use it as a sole carbon source. Ochratoxin α was confirmed as a degradation product in the intracellular extract of A. faecalis using UPLC-MS/MS. Our results suggest that the biodegradation of ochratoxin A by the A. faecalis strain occurs through the hydrolysis of the ochratoxin A amide bond by a putative peptidase. This is the first report to date on the degradation of ochratoxin A by A. faecalis. The A. faecalis strain is presumably a suitable candidate for use in the biodegradation of ochratoxin A. Ochratoxin A, which is produced by some filamentous fungi, severely impacts human and animal health by contaminating several types of food and feed. Our study contributes to the identification of the function of A. faecalis 0D-1, which is capable of producing hydrolytic enzyme(s) to biodegrade ochratoxin A into nontoxic ochratoxin α, to minimize the risk associated with ochratoxin A exposure. © 2017 The Society for Applied Microbiology.

Metabolism of ochratoxin A by rats.

PubMed Central

Støren, O; Holm, H; Størmer, F C

1982-01-01

Albino rats were given ochratoxin A (6.6 mg/kg body weight) intraperitoneally or per os. Independent of route administration, 6% of a given dose was excreted as the toxin, 1 to 1.5% as (4R)-4-hydroxyochratoxin A, and 25 to 27% as ochratoxin alpha in the urine. The metabolite (4S)-4-hydroxyochratoxin A, which is formed by rat liver microsomes in the presence of NADPH, was not detected. Only traces of ochratoxins A and alpha were found in feces. Identical experiments were carried out with brown rats, since the Km value for the formation of the 4S epimer was considerably lower when brown rat microsomes were used. About the same ratios of metabolites and metabolite recoveries as those found for albino rats were found for brown rats. Brown rats were also given the two hydroxylated metabolites and ochratoxin alpha (0.66 mg/kg body weight) intraperitoneally. The three compounds were excreted in the urine; within 48 h, 90% recovery of ochratoxin alpha and 54 and 35%, respectively, of the 4R and 4S isomers were observed. PMID:7149712

The ergot alkaloid gene cluster: functional analyses and evolutionary aspects.

PubMed

Lorenz, Nicole; Haarmann, Thomas; Pazoutová, Sylvie; Jung, Manfred; Tudzynski, Paul

2009-01-01

Ergot alkaloids and their derivatives have been traditionally used as therapeutic agents in migraine, blood pressure regulation and help in childbirth and abortion. Their production in submerse culture is a long established biotechnological process. Ergot alkaloids are produced mainly by members of the genus Claviceps, with Claviceps purpurea as best investigated species concerning the biochemistry of ergot alkaloid synthesis (EAS). Genes encoding enzymes involved in EAS have been shown to be clustered; functional analyses of EAS cluster genes have allowed to assign specific functions to several gene products. Various Claviceps species differ with respect to their host specificity and their alkaloid content; comparison of the ergot alkaloid clusters in these species (and of clavine alkaloid clusters in other genera) yields interesting insights into the evolution of cluster structure. This review focuses on recently published and also yet unpublished data on the structure and evolution of the EAS gene cluster and on the function and regulation of cluster genes. These analyses have also significant biotechnological implications: the characterization of non-ribosomal peptide synthetases (NRPS) involved in the synthesis of the peptide moiety of ergopeptines opened interesting perspectives for the synthesis of ergot alkaloids; on the other hand, defined mutants could be generated producing interesting intermediates or only single peptide alkaloids (instead of the alkaloid mixtures usually produced by industrial strains).

Ergot alkaloids in Norwegian wild grasses: a mass spectrometric approach.

PubMed

Uhlig, Silvio; Vikøren, Turid; Ivanova, Lada; Handeland, Kjell

2007-01-01

Ergot alkaloids are mycotoxins which are produced among fungi in the family Clavicipitaceae. Poisoning with ergot alkaloids is an important veterinary problem in animal husbandry and has recently also been recognised in wild animals. While the poisoning syndrome observed in domestic animals such as cattle, horses and sheep is usually caused by endophyte-infected grass, the recently observed ergotism among Norwegian cervids is probably due to infection of wild grasses with Claviceps. Mass spectrometry is today the method of choice for the rapid qualitative and quantitative determination of many natural compounds. This study uses tandem quadrupole mass spectrometry as well as ion trap mass spectrometry in connection with electrospray(+) ionisation for the quantification, screening and fragmentation of ergot alkaloids in extracts from Claviceps sclerotia that had been picked from wild grasses from several locations in Norway. Ergotamine, ergovaline, ergonovine and ergocryptine were available as standards and were quantified in the extracts, while ergocrystine, ergocornine, ergonine/ergosine, lysergic acid and lysergol were identified on the basis of their molecular weights and semi-quantified. Ergocrystine dominated the alkaloid spectrum of most extracts. Levels of the quantified alkaloids were in the range 0.2-9300 microg/g. Several unknown ergot alkaloids were found in the extracts. MS(n) experiments identified some as simple lysergic acid amide derivatives, while othes are probably related to ergocrystine and ergocryptine by dehydration, dehydrogenation and/or amino acid substitution at R(1) of the peptide moiety.

Diversification of Ergot Alkaloids in Natural and Modified Fungi

PubMed Central

Robinson, Sarah L.; Panaccione, Daniel G.

2015-01-01

Several fungi in two different families––the Clavicipitaceae and the Trichocomaceae––produce different profiles of ergot alkaloids, many of which are important in agriculture and medicine. All ergot alkaloid producers share early steps before their pathways diverge to produce different end products. EasA, an oxidoreductase of the old yellow enzyme class, has alternate activities in different fungi resulting in branching of the pathway. Enzymes beyond the branch point differ among lineages. In the Clavicipitaceae, diversity is generated by the presence or absence and activities of lysergyl peptide synthetases, which interact to make lysergic acid amides and ergopeptines. The range of ergopeptines in a fungus may be controlled by the presence of multiple peptide synthetases as well as by the specificity of individual peptide synthetase domains. In the Trichocomaceae, diversity is generated by the presence or absence of the prenyl transferase encoded by easL (also called fgaPT1). Moreover, relaxed specificity of EasL appears to contribute to ergot alkaloid diversification. The profile of ergot alkaloids observed within a fungus also is affected by a delayed flux of intermediates through the pathway, which results in an accumulation of intermediates or early pathway byproducts to concentrations comparable to that of the pathway end product. PMID:25609183

One hundred million year old ergot: psychotropic compounds in the Cretaceous?

USDA-ARS?s Scientific Manuscript database

A fungal sclerotium similar to sclerotia of the genus Claviceps, commonly known as ergot, was found infecting a grass kernel in Early Cretaceous Myanmar amber. This represents the first fossil record of ergot dating as far back as the Cretaceous period. The fungus, described as Palaeoclaviceps para...

The history of ergot of rye (Claviceps purpurea) II: 1900-1940.

PubMed

Lee, M R

2009-12-01

Ergot, in 1900, was a 'chemical mess'. Henry Wellcome, the pharmaceutical manufacturer, invited Henry Hallett Dale, a physiologist, to join his research department and solve this problem. Dale, in turn, recruited an outstanding group of scientists, including George Barger, Arthur Ewins and Harold Dudley, who would make distinguished contributions not only to the chemistry of ergot but also to the identification of acetylcholine, histamine and tyramine and to studies on their physiological effects. Initially Barger and Dale isolated the compound ergotoxine, but this proved to be a false lead; it was later shown to be a mixture of three different ergot alkaloids. The major success of the Wellcome group was the discovery and isolation of ergometrine, which would prove to be life-saving in postpartum haemorrhage. In 1917 Arthur Stoll and his colleagues started work on ergot at Sandoz Pharmaceuticals in Basel. A series of important results emerged over the next 30 years, including the isolation of ergotamine in 1918, an effective treatment for migraine with aura.

Acute Toxicity of Ochratoxins A and B in Chicks 1

PubMed Central

Peckham, John C.; Doupnik, Ben; Jones, Oscar H.

1971-01-01

Ochratoxins A and B were given to 1-day-old Babcock B-300 cockerels to evaluate acute toxic effects. Two trials with ochratoxin A gave 7-day oral median lethal dose estimates of 116 μg (3.3 mg/kg) and 135 μg (3.9 mg/kg) per chick. Chicks given daily oral doses of 100 μg of ochratoxin A died on the second day. Single subcutaneous doses of 400 μg of ochratoxin A were also lethal. The 7-day oral median lethal dose of B was estimated at 1,890 μg (54 mg/kg) per chick. Chicks given oral doses of 100 μg of ochratoxin B daily for 10 days survived. Sublethal doses of both ochratoxins A and B resulted in growth suppression which was proportional to the amount of ochratoxin given. Visceral gout was the principal gross finding. Microscopic examinations revealed acute nephrosis, hepatic degeneration or focal necrosis, and enteritis. Suppression of hematopoiesis in the bone marrow and depletion of lymphoid elements from the spleen and bursa of Fabricius were frequently seen. Both ochratoxins appeared to have similar pathological effects. This is the first report on the toxicity of ochratoxin B. PMID:4928604

A European Database of Fusarium graminearum and F. culmorum Trichothecene Genotypes

PubMed Central

Pasquali, Matias; Beyer, Marco; Logrieco, Antonio; Audenaert, Kris; Balmas, Virgilio; Basler, Ryan; Boutigny, Anne-Laure; Chrpová, Jana; Czembor, Elżbieta; Gagkaeva, Tatiana; González-Jaén, María T.; Hofgaard, Ingerd S.; Köycü, Nagehan D.; Hoffmann, Lucien; Lević, Jelena; Marin, Patricia; Miedaner, Thomas; Migheli, Quirico; Moretti, Antonio; Müller, Marina E. H.; Munaut, Françoise; Parikka, Päivi; Pallez-Barthel, Marine; Piec, Jonathan; Scauflaire, Jonathan; Scherm, Barbara; Stanković, Slavica; Thrane, Ulf; Uhlig, Silvio; Vanheule, Adriaan; Yli-Mattila, Tapani; Vogelgsang, Susanne

2016-01-01

Fusarium species, particularly Fusarium graminearum and F. culmorum, are the main cause of trichothecene type B contamination in cereals. Data on the distribution of Fusarium trichothecene genotypes in cereals in Europe are scattered in time and space. Furthermore, a common core set of related variables (sampling method, host cultivar, previous crop, etc.) that would allow more effective analysis of factors influencing the spatial and temporal population distribution, is lacking. Consequently, based on the available data, it is difficult to identify factors influencing chemotype distribution and spread at the European level. Here we describe the results of a collaborative integrated work which aims (1) to characterize the trichothecene genotypes of strains from three Fusarium species, collected over the period 2000–2013 and (2) to enhance the standardization of epidemiological data collection. Information on host plant, country of origin, sampling location, year of sampling and previous crop of 1147 F. graminearum, 479 F. culmorum, and 3 F. cortaderiae strains obtained from 17 European countries was compiled and a map of trichothecene type B genotype distribution was plotted for each species. All information on the strains was collected in a freely accessible and updatable database (www.catalogueeu.luxmcc.lu), which will serve as a starting point for epidemiological analysis of potential spatial and temporal trichothecene genotype shifts in Europe. The analysis of the currently available European dataset showed that in F. graminearum, the predominant genotype was 15-acetyldeoxynivalenol (15-ADON) (82.9%), followed by 3-acetyldeoxynivalenol (3-ADON) (13.6%), and nivalenol (NIV) (3.5%). In F. culmorum, the prevalent genotype was 3-ADON (59.9%), while the NIV genotype accounted for the remaining 40.1%. Both, geographical and temporal patterns of trichothecene genotypes distribution were identified. PMID:27092107

Parasitic fungus Claviceps as a source for biotechnological production of ergot alkaloids.

PubMed

Hulvová, Helena; Galuszka, Petr; Frébortová, Jitka; Frébort, Ivo

2013-01-01

Ergot alkaloids produced by the fungus Claviceps parasitizing on cereals, include three major groups: clavine alkaloids, d-lysergic acid and its derivatives and ergopeptines. These alkaloids are important substances for the pharmatech industry, where they are used for production of anti-migraine drugs, uterotonics, prolactin inhibitors, anti-Parkinson agents, etc. Production of ergot alkaloids is based either on traditional field cultivation of ergot-infected rye or on submerged cultures of the fungus in industrial fermentation plants. In 2010, the total production of these alkaloids in the world was about 20,000 kg, of which field cultivation contributed about 50%. This review covers the recent advances in understanding of the genetics and regulation of biosynthesis of ergot alkaloids, focusing on possible applications of the new knowledge to improve the production yield. Copyright © 2012 Elsevier Inc. All rights reserved.

Detection of airborne Stachybotrys chartarum macrocyclic trichothecene mycotoxins in the indoor environment.

PubMed

Brasel, T L; Martin, J M; Carriker, C G; Wilson, S C; Straus, D C

2005-11-01

The existence of airborne mycotoxins in mold-contaminated buildings has long been hypothesized to be a potential occupant health risk. However, little work has been done to demonstrate the presence of these compounds in such environments. The presence of airborne macrocyclic trichothecene mycotoxins in indoor environments with known Stachybotrys chartarum contamination was therefore investigated. In seven buildings, air was collected using a high-volume liquid impaction bioaerosol sampler (SpinCon PAS 450-10) under static or disturbed conditions. An additional building was sampled using an Andersen GPS-1 PUF sampler modified to separate and collect particulates smaller than conidia. Four control buildings (i.e., no detectable S. chartarum growth or history of water damage) and outdoor air were also tested. Samples were analyzed using a macrocyclic trichothecene-specific enzyme-linked immunosorbent assay (ELISA). ELISA specificity was tested using phosphate-buffered saline extracts of the fungal genera Aspergillus, Chaetomium, Cladosporium, Fusarium, Memnoniella, Penicillium, Rhizopus, and Trichoderma, five Stachybotrys strains, and the indoor air allergens Can f 1, Der p 1, and Fel d 1. For test buildings, the results showed that detectable toxin concentrations increased with the sampling time and short periods of air disturbance. Trichothecene values ranged from <10 to >1,300 pg/m3 of sampled air. The control environments demonstrated statistically significantly (P < 0.001) lower levels of airborne trichothecenes. ELISA specificity experiments demonstrated a high specificity for the trichothecene-producing strain of S. chartarum. Our data indicate that airborne macrocyclic trichothecenes can exist in Stachybotrys-contaminated buildings, and this should be taken into consideration in future indoor air quality investigations.

Trichothecene-Genotypes Play a Role in Fusarium Head Blight Disease Spread and Trichothecene Accumulation in Wheat

USDA-ARS?s Scientific Manuscript database

In the current study, we evaluated the impact of the observed North American evolutionary shift in the Fusarium graminearum complex on disease spread, kernel damage, and trichothecene accumulation in resistant and susceptible wheat genotypes. Four inocula were prepared using composites of F. gramin...

Trichothecene mycotoxins and their determinants in settled dust related to grain production.

PubMed

Nordby, Karl-Christian; Halstensen, Anne Straumfors; Elen, Oleif; Clasen, Per-Erik; Langseth, Wenche; Kristensen, Petter; Eduard, Wijnand

2004-01-01

We hypothesise that inhalant exposure to mycotoxins causes developmental outcomes and certain hormone-related cancers that are associated with grain farming in an epidemiological study. The aim of the present study was to identify and validate determinants of measured trichothecene mycotoxins in grain dust as work environmental trichothecene exposure indicators. Settled grain dust was collected in 92 Norwegian farms during seasons of 1999 and 2000. Production characteristics and climatic data were studied as determinants of trichothecenes in settled dust samples obtained during the production of barley (N = 59), oats (N = 32), and spring wheat (N = 13). Median concentrations of trichothecenes in grain dust were <20, 54, and < 50 mg/kg (ranges < 20-340, < 30-2400, and < 50-1200) for deoxynivalenol (DON), HT-2 toxin (HT-2) and T-2 toxin (T-2) respectively. Late blight potato rot (fungal) forecasts have been broadcast in Norway to help prevent this potato disease. Fungal forecasts representing wet, temperate, and humid meteorological conditions were identified as strong determinants of trichothecene mycotoxins in settled grain dust in this study. Differences in cereal species, production properties and districts contributed less to explain mycotoxin concentrations. Fungal forecasts are validated as indicators of mycotoxin exposure of grain farmers and their use in epidemiological studies may be warranted.

Evolution of structural diversity of trichothecenes, a family of toxins produced by plant pathogenic and entomopathogenic fungi.

PubMed

Proctor, Robert H; McCormick, Susan P; Kim, Hye-Seon; Cardoza, Rosa E; Stanley, April M; Lindo, Laura; Kelly, Amy; Brown, Daren W; Lee, Theresa; Vaughan, Martha M; Alexander, Nancy J; Busman, Mark; Gutiérrez, Santiago

2018-04-01

Trichothecenes are a family of terpenoid toxins produced by multiple genera of fungi, including plant and insect pathogens. Some trichothecenes produced by the fungus Fusarium are among the mycotoxins of greatest concern to food and feed safety because of their toxicity and frequent occurrence in cereal crops, and trichothecene production contributes to pathogenesis of some Fusarium species on plants. Collectively, fungi produce over 150 trichothecene analogs: i.e., molecules that share the same core structure but differ in patterns of substituents attached to the core structure. Here, we carried out genomic, phylogenetic, gene-function, and analytical chemistry studies of strains from nine fungal genera to identify genetic variation responsible for trichothecene structural diversity and to gain insight into evolutionary processes that have contributed to the variation. The results indicate that structural diversity has resulted from gain, loss, and functional changes of trichothecene biosynthetic (TRI) genes. The results also indicate that the presence of some substituents has arisen independently in different fungi by gain of different genes with the same function. Variation in TRI gene duplication and number of TRI loci was also observed among the fungi examined, but there was no evidence that such genetic differences have contributed to trichothecene structural variation. We also inferred ancestral states of the TRI cluster and trichothecene biosynthetic pathway, and proposed scenarios for changes in trichothecene structures during divergence of TRI cluster homologs. Together, our findings provide insight into evolutionary processes responsible for structural diversification of toxins produced by pathogenic fungi.

Faces of a changing climate: semi-quantitative multi-mycotoxin analysis of grain grown in exceptional climatic conditions in Norway.

PubMed

Uhlig, Silvio; Eriksen, Gunnar Sundstøl; Hofgaard, Ingerd Skow; Krska, Rudolf; Beltrán, Eduardo; Sulyok, Michael

2013-09-27

Recent climatological research predicts a significantly wetter climate in Southern Norway as a result of global warming. Thus, the country has already experienced unusually wet summer seasons in the last three years (2010-2012). The aim of this pilot study was to apply an existing multi-analyte LC-MS/MS method for the semi-quantitative determination of 320 fungal and bacterial metabolites in Norwegian cereal grain samples from the 2011 growing season. Such knowledge could provide important information for future survey and research programmes in Norway. The method includes all regulated and well-known mycotoxins such as aflatoxins, trichothecenes, ochratoxin A, fumonisins and zearalenone. In addition, a wide range of less studied compounds are included in the method, e.g., Alternaria toxins, ergot alkaloids and other metabolites produced by fungal species within Fusarium, Penicillium and Aspergillus. Altogether, 46 metabolites, all of fungal origin, were detected in the 76 barley, oats and wheat samples. The analyses confirmed the high prevalence and relatively high concentrations of type-A and -B trichothecenes (e.g., deoxynivalenol up to 7230 µg/kg, HT-2 toxin up to 333 µg/kg). Zearalenone was also among the major mycotoxins detected (maximum concentration 1670 µg/kg). Notably, several other Fusarium metabolites such as culmorin, 2-amino-14,16-dimethyloctadecan-3-ol and avenacein Y were co-occurring. Furthermore, the most prevalent Alternaria toxin was alternariol with a maximum concentration of 449 µg/kg. A number of Penicillium and Aspergillus metabolites were also detected in the samples, e.g., sterigmatocystin in concentrations up to 20 µg/kg.

Recent investigations of ergot alkaloids incorporated into plant and/or animal systems

USDA-ARS?s Scientific Manuscript database

Ergot alkaloids produced by fungi have a basic chemical structure but different chemical moieties at substituent sites resulting in various forms of alkaloids that are distinguishable from one another. Since the ergoline ring structure found in ergot alkaloids is similar to that of biogenic amines (...

Ergot cluster-encoded catalase is required for synthesis of chanoclavine-I in Aspergillus fumigatus.

PubMed

Goetz, Kerry E; Coyle, Christine M; Cheng, Johnathan Z; O'Connor, Sarah E; Panaccione, Daniel G

2011-06-01

Genes required for ergot alkaloid biosynthesis are clustered in the genomes of several fungi. Several conserved ergot cluster genes have been hypothesized, and in some cases demonstrated, to encode early steps of the pathway shared among fungi that ultimately make different ergot alkaloid end products. The deduced amino acid sequence of one of these conserved genes (easC) indicates a catalase as the product, but a role for a catalase in the ergot alkaloid pathway has not been established. We disrupted easC of Aspergillus fumigatus by homologous recombination with a truncated copy of that gene. The resulting mutant (ΔeasC) failed to produce the ergot alkaloids typically observed in A. fumigatus, including chanoclavine-I, festuclavine, and fumigaclavines B, A, and C. The ΔeasC mutant instead accumulated N-methyl-4-dimethylallyltryptophan (N-Me-DMAT), an intermediate recently shown to accumulate in Claviceps purpurea strains mutated at ccsA (called easE in A. fumigatus) (Lorenz et al. Appl Environ Microbiol 76:1822-1830, 2010). A ΔeasE disruption mutant of A. fumigatus also failed to accumulate chanoclavine-I and downstream ergot alkaloids and, instead, accumulated N-Me-DMAT. Feeding chanoclavine-I to the ΔeasC mutant restored ergot alkaloid production. Complementation of either ΔeasC or ΔeasE mutants with the respective wild-type allele also restored ergot alkaloid production. The easC gene was expressed in Escherichia coli, and the protein product displayed in vitro catalase activity with H(2)O(2) but did not act, in isolation, on N-Me-DMAT as substrate. The data indicate that the products of both easC (catalase) and easE (FAD-dependent oxidoreductase) are required for conversion of N-Me-DMAT to chanoclavine-I.

L-Threonine and its analogue added to autoclaved solid medium suppress trichothecene production by Fusarium graminearum.

PubMed

Maeda, Kazuyuki; Nakajima, Yuichi; Tanahashi, Yoshikazu; Kitou, Yoshiyuki; Miwa, Akihiro; Kanamaru, Kyoko; Kobayashi, Tetsuo; Nishiuchi, Takumi; Kimura, Makoto

2017-08-01

Fusarium graminearum produces trichothecene mycotoxins under certain nutritional conditions. When L-Thr and its analogue L-allo-threonine were added to brown rice flour solid medium before inoculation, trichothecene production after 4 days of incubation was suppressed. A time-course analysis of gene expression demonstrated that L-Thr suppressed transcription of Tri6, a trichothecene master regulator gene, and a terpene cyclase Tri5 gene. Regulation of trichothecene biosynthesis by altering major primary metabolic processes may open up the possibility to develop safe chemicals for the reduction of mycotoxin contamination might be developed.

Production of Ochratoxins in Different Cereal Products by Aspergillus ochraceus1

PubMed Central

Trenk, Hugh L.; Butz, Mary E.; Chu, Fun Sun

1971-01-01

The effects of temperature and length of incubation on ochratoxin A production in various substrates were studied. The optimal temperature for toxin production by Aspergillus ochraceus NRRL-3174 was found to be around 28 C. Very low levels of ochratoxin A are produced in corn, rice, and wheat bran at 4 C. The optimal time for ochratoxin A production depends on the substrate, ranging from 7 to 14 days at 28 C. Ochratoxin B and dihydroisocoumaric acid, i.e., one of the hydrolysis products of ochratoxin A, were produced in rice but at levels considerably lower than ochratoxin A. No ochratoxin C was produced in rice at 28 C. When added to rice cereal or oatmeal, the toxin was found to be very stable over prolonged storage and even to autoclaving for 3 hr. PMID:5564676

The Role of Trichothecene-Chemotypes in Fusarium Head Blight Disease Spread and Trichothecene Accumulation in Wheat

USDA-ARS?s Scientific Manuscript database

Three major strain-specific trichothecene-chemotypes have been identified in F. graminearum-infected crops in North America: 3-acetyldeoxynivalenol (3ADON), 15ADON, and nivalenol (NIV). The emergence of the 3ADON- and NIV-chemotypes on the continent is a fairly recent phenomenon. In addition, str...

Relevance of trichothecenes in fungal physiology: Disruption of tri5 in Trichoderma arundinaceum

USDA-ARS?s Scientific Manuscript database

Trichothecenes are sesquiterpenoid mycotoxins produced mainly by Fusarium species. Harzianum A (HA), a non-phytotoxic trichothecene produced by Trichoderma arundinaceum, has recently been found to have antagonistic activity against fungal plant pathogens and to induce plant genes involved in defense...

Biology, Genetics, and Management of Ergot (Claviceps spp.) in Rye, Sorghum, and Pearl Millet

PubMed Central

Miedaner, Thomas; Geiger, Hartwig H.

2015-01-01

Ergot is a disease of cereals and grasses caused by fungi in the genus Claviceps. Of particular concern are Claviceps purpurea in temperate regions, C. africana in sorghum (worldwide), and C. fusiformis in pearl millet (Africa, Asia). The fungi infect young, usually unfertilized ovaries, replacing the seeds by dark mycelial masses known as sclerotia. The percentage of sclerotia in marketable grain is strictly regulated in many countries. In winter rye, ergot has been known in Europe since the early Middle Ages. The alkaloids produced by the fungus severely affect the health of humans and warm-blooded animals. In sorghum and pearl millet, ergot became a problem when growers adopted hybrid technology, which increased host susceptibility. Plant traits reducing ergot infection include immediate pollination of receptive stigmas, closed flowering (cleistogamy), and physiological resistance. Genetic, nonpollen-mediated variation in ergot susceptibility could be demonstrated in all three affected cereals. Fungicides have limited efficacy and application is weather dependent. Sorting out the sclerotia from the harvest by photocells is expensive and time consuming. In conclusion, molecular-based hybrid rye breeding could improve pollen fertility by introgressing effective restorer genes thus bringing down the ergot infection level to that of conventional population cultivars. A further reduction might be feasible in the future by selecting more resistant germplasm. PMID:25723323

Biology, genetics, and management of ergot (Claviceps spp.) in rye, sorghum, and pearl millet.

PubMed

Miedaner, Thomas; Geiger, Hartwig H

2015-02-25

Ergot is a disease of cereals and grasses caused by fungi in the genus Claviceps. Of particular concern are Claviceps purpurea in temperate regions, C. africana in sorghum (worldwide), and C. fusiformis in pearl millet (Africa, Asia). The fungi infect young, usually unfertilized ovaries, replacing the seeds by dark mycelial masses known as sclerotia. The percentage of sclerotia in marketable grain is strictly regulated in many countries. In winter rye, ergot has been known in Europe since the early Middle Ages. The alkaloids produced by the fungus severely affect the health of humans and warm-blooded animals. In sorghum and pearl millet, ergot became a problem when growers adopted hybrid technology, which increased host susceptibility. Plant traits reducing ergot infection include immediate pollination of receptive stigmas, closed flowering (cleistogamy), and physiological resistance. Genetic, nonpollen-mediated variation in ergot susceptibility could be demonstrated in all three affected cereals. Fungicides have limited efficacy and application is weather dependent. Sorting out the sclerotia from the harvest by photocells is expensive and time consuming. In conclusion, molecular-based hybrid rye breeding could improve pollen fertility by introgressing effective restorer genes thus bringing down the ergot infection level to that of conventional population cultivars. A further reduction might be feasible in the future by selecting more resistant germplasm.

Trichothecenes NIV and DON modulate the maturation of murine dendritic cells.

PubMed

Luongo, D; Severino, L; Bergamo, P; D'Arienzo, R; Rossi, M

2010-01-01

Nivalenol (NIV) and Deoxynivalenol (DON), mycotoxins of the trichothecene family are considered very common food contaminants. In this work, we investigated whether the immunotoxic effects ascribed to these trichothecenes may be mediated by perturbations in the activity of dendritic cells (DCs). Murine bone marrow-derived DCs were used to evaluate the effects of NIV and DON on the LPS-induced maturation process. We found that the expression of the class II MHC and of the accessory CD11c molecules, but not of the costimulatory CD86 marker, was down-regulated by NIV and DON exposure in LPS-treated DCs, as well as nitric oxide (NO) production. Interestingly, NIV, but not DON, induced DC necrosis. Moreover, the analysis of the cytokine pattern showed that IL-12 and IL-10 expressions induced by LPS exposure were suppressed by both trichothecenes in a dose-dependent fashion. On the other hand, the secretion of the proinflammatory cytokine TNF-alpha was increased as a direct consequence of DON and NIV exposure. Taken together, our data indicated that the immunotoxicity of NIV and DON was related to the capacity of both trichothecenes to interfere with phenotypic and functional features of maturing DCs.

Evolution of structural diversity of trichothecenes, a family of toxins produced by plant pathogenic and entomopathogenic fungi

PubMed Central

McCormick, Susan P.; Lee, Theresa; Vaughan, Martha M.; Alexander, Nancy J.; Busman, Mark

2018-01-01

Trichothecenes are a family of terpenoid toxins produced by multiple genera of fungi, including plant and insect pathogens. Some trichothecenes produced by the fungus Fusarium are among the mycotoxins of greatest concern to food and feed safety because of their toxicity and frequent occurrence in cereal crops, and trichothecene production contributes to pathogenesis of some Fusarium species on plants. Collectively, fungi produce over 150 trichothecene analogs: i.e., molecules that share the same core structure but differ in patterns of substituents attached to the core structure. Here, we carried out genomic, phylogenetic, gene-function, and analytical chemistry studies of strains from nine fungal genera to identify genetic variation responsible for trichothecene structural diversity and to gain insight into evolutionary processes that have contributed to the variation. The results indicate that structural diversity has resulted from gain, loss, and functional changes of trichothecene biosynthetic (TRI) genes. The results also indicate that the presence of some substituents has arisen independently in different fungi by gain of different genes with the same function. Variation in TRI gene duplication and number of TRI loci was also observed among the fungi examined, but there was no evidence that such genetic differences have contributed to trichothecene structural variation. We also inferred ancestral states of the TRI cluster and trichothecene biosynthetic pathway, and proposed scenarios for changes in trichothecene structures during divergence of TRI cluster homologs. Together, our findings provide insight into evolutionary processes responsible for structural diversification of toxins produced by pathogenic fungi. PMID:29649280

Ergot Alkaloids (Re)generate New Leads as Antiparasitics

PubMed Central

Chan, John D.; Agbedanu, Prince N.; Grab, Thomas; Zamanian, Mostafa; Dosa, Peter I.; Day, Timothy A.; Marchant, Jonathan S.

2015-01-01

Abstract Praziquantel (PZQ) is a key therapy for treatment of parasitic flatworm infections of humans and livestock, but the mechanism of action of this drug is unresolved. Resolving PZQ-engaged targets and effectors is important for identifying new druggable pathways that may yield novel antiparasitic agents. Here we use functional, genetic and pharmacological approaches to reveal that serotonergic signals antagonize PZQ action in vivo. Exogenous 5-hydroxytryptamine (5-HT) rescued PZQ-evoked polarity and mobility defects in free-living planarian flatworms. In contrast, knockdown of a prevalently expressed planarian 5-HT receptor potentiated or phenocopied PZQ action in different functional assays. Subsequent screening of serotonergic ligands revealed that several ergot alkaloids possessed broad efficacy at modulating regenerative outcomes and the mobility of both free living and parasitic flatworms. Ergot alkaloids that phenocopied PZQ in regenerative assays to cause bipolar regeneration exhibited structural modifications consistent with serotonergic blockade. These data suggest that serotonergic activation blocks PZQ action in vivo, while serotonergic antagonists phenocopy PZQ action. Importantly these studies identify the ergot alkaloid scaffold as a promising structural framework for designing potent agents targeting parasitic bioaminergic G protein coupled receptors. PMID:26367744

Ochratoxin A in cereal-based baby foods: occurrence and safety evaluation.

PubMed

Beretta, B; De Domenico, R; Gaiaschi, A; Ballabio, C; Galli, C L; Gigliotti, C; Restani, P

2002-01-01

Ochratoxin A is a typical cereal contaminant with strong nephrotoxic activity. To estimate the quantity of ochratoxin A that can be taken in by a child in the weaning period, several samples of cereal-based baby foods were analysed. Although most samples analysed contained ochratoxin A in undetectable amounts or below the Italian legal limit of 0.5 microg kg(-1), some irregular products were found. In particular, the analyses of the 119 batches (338 samples) of baby foods considered indicated that: 20 batches (16.8%) contained detectable quantities of ochratoxin A and four of these (3.4% of the total) contained ochratoxin A above the Italian permitted value. All samples coming from agricultural practices based on integrated pest management contained undetectable amounts of ochratoxin A, while approximately 5% of batches coming from conventional and organic agricultural practices were above the legal limit. On the basis of the established provisional tolerable weekly intake (PTWI), there is no significant toxicological risk for a child who occasionally consumes a formula with ochratoxin concentration slightly above the permitted level. However, stricter controls have to be applied to reject the batches containing irregular concentrations of ochratoxin A.

Evidence for an ergot alkaloid gene cluster in Claviceps purpurea.

PubMed

Tudzynski, P; Hölter, K; Correia, T; Arntz, C; Grammel, N; Keller, U

1999-02-01

A gene (cpd1) coding for the dimethylallyltryptophan synthase (DMATS) that catalyzes the first specific step in the biosynthesis of ergot alkaloids, was cloned from a strain of Claviceps purpurea that produces alkaloids in axenic culture. The derived gene product (CPD1) shows only 70% similarity to the corresponding gene previously isolated from Claviceps strain ATCC 26245, which is likely to be an isolate of C. fusiformis. Therefore, the related cpd1 most probably represents the first C. purpurea gene coding for an enzymatic step of the alkaloid biosynthetic pathway to be cloned. Analysis of the 3'-flanking region of cpd1 revealed a second, closely linked ergot alkaloid biosynthetic gene named cpps1, which codes for a 356-kDa polypeptide showing significant similarity to fungal modular peptide synthetases. The protein contains three amino acid-activating modules, and in the second module a sequence is found which matches that of an internal peptide (17 amino acids in length) obtained from a tryptic digest of lysergyl peptide synthetase 1 (LPS1) of C. purpurea, thus confirming that cpps1 encodes LPS1. LPS1 activates the three amino acids of the peptide portion of ergot peptide alkaloids during D-lysergyl peptide assembly. Chromosome walking revealed the presence of additional genes upstream of cpd1 which are probably also involved in ergot alkaloid biosynthesis: cpox1 probably codes for an FAD-dependent oxidoreductase (which could represent the chanoclavine cyclase), and a second putative oxidoreductase gene, cpox2, is closely linked to it in inverse orientation. RT-PCR experiments confirm that all four genes are expressed under conditions of peptide alkaloid biosynthesis. These results strongly suggest that at least some genes of ergot alkaloid biosynthesis in C. purpurea are clustered, opening the way for a detailed molecular genetic analysis of the pathway.

Sorption of Ochratoxin A from Aqueous Solutions Using β-Cyclodextrin-Polyurethane Polymer

PubMed Central

Appell, Michael; Jackson, Michael A.

2012-01-01

The ability of a cyclodextrin-polyurethane polymer to remove ochratoxin A from aqueous solutions was examined by batch rebinding assays. The results from the aqueous binding studies were fit to two parameter models to gain insight into the interaction of ochratoxin A with the nanosponge material. The ochratoxin A sorption data fit well to the heterogeneous Freundlich isotherm model. The polymer was less effective at binding ochratoxin A in high pH buffer (9.5) under conditions where ochratoxin A exists predominantly in the dianionic state. Batch rebinding assays in red wine indicate the polymer is able to remove significant levels of ochratoxin A from spiked solutions between 1–10 μg·L−1. These results suggest cyclodextrin nanosponge materials are suitable to reduce levels of ochratoxin A from spiked aqueous solutions and red wine samples. PMID:22474569

Sorption of ochratoxin A from aqueous solutions using β-cyclodextrin-polyurethane polymer.

PubMed

Appell, Michael; Jackson, Michael A

2012-02-01

The ability of a cyclodextrin-polyurethane polymer to remove ochratoxin A from aqueous solutions was examined by batch rebinding assays. The results from the aqueous binding studies were fit to two parameter models to gain insight into the interaction of ochratoxin A with the nanosponge material. The ochratoxin A sorption data fit well to the heterogeneous Freundlich isotherm model. The polymer was less effective at binding ochratoxin A in high pH buffer (9.5) under conditions where ochratoxin A exists predominantly in the dianionic state. Batch rebinding assays in red wine indicate the polymer is able to remove significant levels of ochratoxin A from spiked solutions between 1-10 μg·L(-1). These results suggest cyclodextrin nanosponge materials are suitable to reduce levels of ochratoxin A from spiked aqueous solutions and red wine samples.

Detection of Total Ergot Alkaloids in Cereal Flour and in Bread by a Generic Enzyme Immunoassay Method.

PubMed

Gross, Madeleine; Curtui, Valeriu; Usleber, Ewald

2018-05-01

Four sets of polyclonal antibodies against ergot alkaloids ergometrine, ergotamine, α-ergocryptine, and ergocornine were produced and characterized in a competitive direct or indirect enzyme immunoassay (EIA). Standard curve LODs were 0.03 ng/mL (ergometrine EIA) to 2.0 ng/mL (ergocornine EIA). Three EIAs were highly specific, whereas the ergometrine EIA had a broad specificity pattern and reacted, albeit weakly, with all seven major ergot alkaloids and their epimeric forms. Using the ergometrine EIA, a generic test system was established in which total ergot alkaloids are quantified by a standard curve for a toxin mixture composed of three alkaloids that matched the ergot alkaloid composition in naturally contaminated rye and wheat products. Sample extraction with acetonitrile-phosphate-buffered saline at pH 6.0 without further cleanup was sufficient for EIA analysis. The LODs for total ergot alkaloids were 20 ng/g in rye and wheat flour and 14 ng/g in bread. Recoveries were 85-110% (RSDs of 0.1-11.7%) at a concentration range of 50-1000 ng/g. The total ergot alkaloid EIA was validated by comparison with HPLC-fluorescence detection. Although some under- and overestimation by the total ergot alkaloid EIA was observed, it was suitable for the reliable identification of positive samples at 10-20 ng/g and for the determination of total ergot alkaloids in a concentration range between 100 and 1000 ng/g.

Sorghum ergot (Claviceps africana) associated with agalactia and feed refusal in pigs and dairy cattle.

PubMed

Blaney, B J; McKenzie, R A; Walters, J R; Taylor, L F; Bewg, W S; Ryley, M J; Maryam, R

2000-02-01

To establish the aetiology and define the main clinical features of a syndrome characterised by severe feed refusal, death of piglets and reduced milk production in pigs and dairy cattle. Clinical, pathological, toxicological and epidemiological examination of clinical cases in 10 piggeries and 4 dairies, located between 50 and 150 km south-west of Rockhampton. All cases were associated with the feeding of sorghum grain infected with sorghum ergot (Claviceps africana). There was mild to severe feed refusal when the sorghum was first offered. Sows fed ergot before farrowing had shrunken udders, produced no colostrum, and displayed signs of oestrus. All of their piglets died, apparently from starvation: necropsy of a few piglets showed that they were born alive and walked, but had ingested no milk. Sows fed the grain after farrowing had severe reductions in milk production despite aggressive sucking by piglets, leading to very poor growth of piglets. There were no signs of infectious disease. Ergot in sorghum samples ranged from 1 to 31% ergot sclerotes by weight. Total alkaloid concentrations in mixed feeds ranged from 5 to 40 mg/kg, with dihydroergosine accounting for approximately 90%. At the same time, in the same districts, there were reports of feed refusal and reduced milk production from 4 dairy farms. Grain samples from these farms contained up to 17% C africana ergot sclerotia. Agalactia and feed refusal are classical signs of poisoning by rye ergot (C purpurea), but this is the first time that sorghum ergot has been associated with a similar syndrome.

Online detection and quantification of ergot bodies in cereals using near infrared hyperspectral imaging.

PubMed

Vermeulen, Ph; Pierna, J A Fernández; Egmond, H P van; Dardenne, P; Baeten, V

2012-01-01

The occurrence of ergot bodies (sclerotia of Claviceps purpurea) in cereals presents a high toxicity risk for animals and humans due to the alkaloid content. To reduce this risk, the European Commission fixed an ergot concentration limit of 0.1% in all feedstuffs containing unground cereals, and a limit of 0.05% in 'intervention' cereals destined for humans. This study sought to develop a procedure based on near infrared hyperspectral imaging and multivariate image analysis to detect and quantify ergot contamination in cereals. Hyperspectral images were collected using an NIR hyperspectral line scan combined with a conveyor belt. All images consisted of lines of 320 pixels that were acquired at 209 wavelength channels (1100-2400 nm). To test the procedure, several wheat samples with different levels of ergot contamination were prepared. The results showed a correlation higher than 0.99 between the predicted values obtained using chemometric tools such as partial least squares discriminant analysis or support vector machine and the reference values. For a wheat sample with a level of ergot contamination as low as 0.01 %, it was possible to identify groups of pixels detected as ergot to conclude that the sample was contaminated. In addition, no false positives were obtained with non-contaminated samples. The limit of detection was found to be 145 mg/kg and the limit of quantification 341 mg/kg. The reproducibility tests of the measurements performed over several weeks showed that the results were always within the limits allowed. Additional studies were done to optimise the parameters in terms of number of samples analysed per unit of time or conveyor belt speed. It was shown that ergot can be detected using a speed of 1-100 mm/s and that a sample of 250 g can be analysed in 1 min.

Chemical, Physical and Biological Approaches to Prevent Ochratoxin Induced Toxicoses in Humans and Animals

PubMed Central

Varga, János; Kocsubé, Sándor; Péteri, Zsanett; Vágvölgyi, Csaba; Tóth, Beáta

2010-01-01

Ochratoxins are polyketide derived fungal secondary metabolites with nephrotoxic, immunosuppressive, teratogenic, and carcinogenic properties. Ochratoxin-producing fungi may contaminate agricultural products in the field (preharvest spoilage), during storage (postharvest spoilage), or during processing. Ochratoxin contamination of foods and feeds poses a serious health hazard to animals and humans. Several strategies have been investigated for lowering the ochratoxin content in agricultural products. These strategies can be classified into three main categories: prevention of ochratoxin contamination, decontamination or detoxification of foods contaminated with ochratoxins, and inhibition of the absorption of consumed ochratoxins in the gastrointestinal tract. This paper gives an overview of the strategies that are promising with regard to lowering the ochratoxin burden of animals and humans. PMID:22069658

Ergot alkaloid intoxication in perennial ryegrass (Lolium perenne): an emerging animal health concern in Ireland?

PubMed

Canty, Mary J; Fogarty, Ursula; Sheridan, Michael K; Ensley, Steve M; Schrunk, Dwayne E; More, Simon J

2014-01-01

Four primary mycotoxicosis have been reported in livestock caused by fungal infections of grasses or cereals by members of the Clavicipitaceae family. Ergotism (generally associated with grasses, rye, triticale and other grains) and fescue toxicosis (associated with tall fescue grass, Festuca arundinacea) are both caused by ergot alkaloids, and referred to as 'ergot alkaloid intoxication'. Ryegrass staggers (associated with perennial ryegrass Lolium perenne) is due to intoxication with an indole-diperpene, Lolitrem B, and metabolites. Fescue-associated oedema, recently described in Australia, may be associated with a pyrrolizidine alkaloid, N-acetyl norloline. Ergotism, caused by the fungus Claviceps purpurea, is visible and infects the outside of the plant seed. Fescue toxicosis and ryegrass staggers are caused by Neotyphodium coenophalium and N. lolii, respectively. Fescue-associated oedema has been associated with tall fescue varieties infected with a specific strain of N. coenophialum (AR542, Max P or Max Q). The name Neotyphodium refers to asexual derivatives of Epichloë spp., which have collectively been termed the epichloë fungi. These fungi exist symbiotically within the grass and are invisible to the naked eye. The primary toxicological effect of ergot alkaloid involves vasoconstriction and/or hypoprolactinaemia. Ingestion of ergot alkaloid by livestock can cause a range of effects, including poor weight gain, reduced fertility, hyperthermia, convulsions, gangrene of the extremities, and death. To date there are no published reports, either internationally or nationally, reporting ergot alkaloid intoxication specifically associated with perennial ryegrass endophytes. However, unpublished reports from the Irish Equine Centre have identified a potential emerging problem of ergot alkaloid intoxication with respect to equines and bovines, on primarily perennial ryegrass-based diets. Ergovaline has been isolated in varying concentrations in the herbage of a

Ergot alkaloid intoxication in perennial ryegrass (Lolium perenne): an emerging animal health concern in Ireland?

PubMed Central

2014-01-01

Four primary mycotoxicosis have been reported in livestock caused by fungal infections of grasses or cereals by members of the Clavicipitaceae family. Ergotism (generally associated with grasses, rye, triticale and other grains) and fescue toxicosis (associated with tall fescue grass, Festuca arundinacea) are both caused by ergot alkaloids, and referred to as ‘ergot alkaloid intoxication’. Ryegrass staggers (associated with perennial ryegrass Lolium perenne) is due to intoxication with an indole-diperpene, Lolitrem B, and metabolites. Fescue-associated oedema, recently described in Australia, may be associated with a pyrrolizidine alkaloid, N-acetyl norloline. Ergotism, caused by the fungus Claviceps purpurea, is visible and infects the outside of the plant seed. Fescue toxicosis and ryegrass staggers are caused by Neotyphodium coenophalium and N. lolii, respectively. Fescue-associated oedema has been associated with tall fescue varieties infected with a specific strain of N. coenophialum (AR542, Max P or Max Q). The name Neotyphodium refers to asexual derivatives of Epichloë spp., which have collectively been termed the epichloë fungi. These fungi exist symbiotically within the grass and are invisible to the naked eye. The primary toxicological effect of ergot alkaloid involves vasoconstriction and/or hypoprolactinaemia. Ingestion of ergot alkaloid by livestock can cause a range of effects, including poor weight gain, reduced fertility, hyperthermia, convulsions, gangrene of the extremities, and death. To date there are no published reports, either internationally or nationally, reporting ergot alkaloid intoxication specifically associated with perennial ryegrass endophytes. However, unpublished reports from the Irish Equine Centre have identified a potential emerging problem of ergot alkaloid intoxication with respect to equines and bovines, on primarily perennial ryegrass-based diets. Ergovaline has been isolated in varying concentrations in the herbage of

Drug dependence associated with triptans and ergot derivatives: a case/non-case study.

PubMed

Beau-Salinas, Frédérique; Jonville-Béra, Annie Pierre; Cissoko, Haware; Bensouda-Grimaldi, Lamiae; Autret-Leca, Elisabeth

2010-04-01

The aim of this case/non-case study was to assess and compare the risk of drug dependence associated with different migraine-specific drugs, i.e., ergot derivatives and triptans, using the French pharmacovigilance database. Reports on drug side effects recorded in this database between January 1985 and June 2007 were analyzed, and triptans (almotriptan, eletriptan, naratriptan, sumatriptan, and zolmitriptan) as well as ergot derivatives used in acute migraine were examined. For all reports, cases were defined as those reports corresponding to "drug abuse," "physical or mental drug dependence," and "pharmacodependence," whereas "non-cases" were defined as all the remaining SED reports. The method's reliability was assessed by calculating the risk associated with a negative (amoxicillin) and a positive (benzodiazepines) control. The risk of dependence associated with each drug and control was evaluated by calculating the odds ratio (OR) with a confidence interval of 95%. Among the 309,178 reports recorded in the database, drug dependence accounted for 0.8% (2,489) of the reports, with 10.9% (449) involving a triptan, and 9.33% (332) an ergot derivative. The risk of dependence was similar for triptans and ergot derivatives and did not differ from that of benzodiazepines. In the triptan group, the risk (odds ratio [95% CI]) ranged from 10.3 [4.8-22.3] for sumatriptan to 21.5 for eletriptan [10.1-45.6], while in the ergot derivative group, it ranged from 12 [8-17.9] for ergotamine to 20.6 [8-53] for dihydroergotamine. These findings confirm the hypothesis that triptans and ergot derivatives are associated with an increased risk of drug dependence.

The AtNFXL1 gene functions as a signaling component of the type A trichothecene-dependent response

PubMed Central

Asano, Tomoya; Yasuda, Michiko; Nakashita, Hideo; Kimura, Makoto; Yamaguchi1, Kazuo

2008-01-01

Phytopathogenic Fusarium species produce the trichothecene family of phytotoxins, which function as a virulence factor during infection of plants. Trichothecenes are classifiable into four major groups by their chemical structures. Recently, the AtNFXL1 gene was reported as a type A trichothecene T-2 toxin-inducible gene. The AtNFXL1 gene encodes a putative transcription factor with similarity to the human transcription repressor NF-X1. The atnfxl1 mutant exhibited hypersensitivity phenotype to T-2 toxin but not to type B deoxynivalenol (DON) in comparison with wild type when Arabidopsis thaliana grew on agar medium containing trichothecenes. The absence or presence of a carbonyl group at the C8 position distinguishes type A and type B. Growth defect by another type A trichothecene diacetoxyscirpenol (DAS), was weakly enhanced in the atnfxl1 mutant. Diacetoxyscirpenol is distinguishable from T-2 toxin only by the absence of an isovaleryl group at the C8 position. Correspondingly, the AtNFXL1 promoter activity was apparently induced in T-2 toxin-treated and DAS-treated plants. In contrast, DON failed to induce the AtNFXL1 promoter activity. Consequently, the AtNFXL1 gene functions as a signaling component of the type A trichothecene-dependent response in Arabidopsis. In addition, the C8 position of trichothecenes might be closely related to the function of AtNFXL1 gene. PMID:19704430

Ergot Alkaloids Produced by Endophytic Fungi of the Genus Epichloë

PubMed Central

Guerre, Philippe

2015-01-01

The development of fungal endophytes of the genus Epichloë in grasses results in the production of different groups of alkaloids, whose mechanism and biological spectrum of toxicity can differ considerably. Ergot alkaloids, when present in endophyte-infected tall fescue, are responsible for “fescue toxicosis” in livestock, whereas indole-diterpene alkaloids, when present in endophyte-infected ryegrass, are responsible for “ryegrass staggers”. In contrast, peramine and loline alkaloids are deterrent and/or toxic to insects. Other toxic effects in livestock associated with the consumption of endophyte-infected grass that contain ergot alkaloids include the “sleepy grass” and “drunken horse grass” diseases. Although ergovaline is the main ergopeptine alkaloid produced in endophyte-infected tall fescue and is recognized as responsible for fescue toxicosis, a number of questions still exist concerning the profile of alkaloid production in tall fescue and the worldwide distribution of tall fescue toxicosis. The purpose of this review is to present ergot alkaloids produced in endophyte-infected grass, the factors of variation of their level in plants, and the diseases observed in the mammalian species as relate to the profiles of alkaloid production. In the final section, interactions between ergot alkaloids and drug-metabolizing enzymes are presented as mechanisms that could contribute to toxicity. PMID:25756954

Blocking aflatoxins in corn by using non-toxigenic strains of Aspergillus flavus

USDA-ARS?s Scientific Manuscript database

There are over 500 previously reported mycotoxins. However, only a few have been identified as important for food safety, including aflatoxins, fumonisins, cyclopiazonic acid (CPA), trichothecenes, zearalenone, ochratoxins, and patulin. Mycotoxins contaminate plant materials, causing acute and ch...

Lactam ergot alkaloids (ergopeptams) as predominant alkaloids in sclerotia of Claviceps purpurea from Norwegian wild grasses.

PubMed

Uhlig, Silvio; Petersen, Dirk

2008-07-01

Four major alkaloids in the extracts from sclerotia of Claviceps purpurea, picked from wild grasses, have been identified as lactam (non-cyclol) ergot alkaloids. The structural information was obtained from ion trap MS and NMR spectroscopy. The data for one of the lactam ergot alkaloids were coinciding with ergocristam [N-(lysergyl-valyl)-cyclo(phenylalanyl-prolyl)]. The structural information of two further lactam alkaloids was suggestive of either alpha- or beta-ergocryptam [N-(lysergyl-valyl)-cyclo(leucyl-prolyl) or N-(lysergyl-valyl)-cyclo(isoleucyl-prolyl)] and ergoannam [N-(lysergyl-leucyl)-cyclo(leucyl-prolyl) or N-(lysergyl-isoleucyl)-cyclo(isoleucyl-prolyl)]. The constitution of the fourth lactam ergot alkaloid corresponded to N-(lysergyl-isoleucyl)-cyclo(phenylalanyl-prolyl), a new ergopeptam, which has not been described before. Additionally, the cyclol-analogue of the new ergopeptam was detected in the extracts and has been identified on the basis of its product ion spectrum from fragmentation of [M+H](+). The study described in this paper shows that lactam ergot alkaloids may not only be minor products of ergopeptine biosynthesis, as has been suggested hitherto, but may be major biosynthetic endproducts for some ergot strains. This is also the first report demonstrating the production of an ergot alkaloid that contains isoleucine as the second amino acid, i.e. the N-(lysergyl-isoleucyl)-moiety, by parasitic, naturally growing C. purpurea. This unusual type of ergot alkaloid has so far only been found in saprophytic cultures of C. purpurea.

Spatial patterns of ergot and quantification of sclerotia in perennial ryegrass seed fields

USDA-ARS?s Scientific Manuscript database

Ergot, caused by Claviceps purpurea, is a major disease of perennial ryegrass grass grown for seed in eastern Oregon. The objective of this research was to quantify and describe the spatial patterns of ergot severity in each of three 50 ha commercial fields of perennial ryegrass grown for seed in 20...

Variation in type A trichothecene production and trichothecene biosynthetic genes in Fusarium goolgardi from natural ecosystems of Australia

USDA-ARS?s Scientific Manuscript database

The fungal species Fusarium goolgardi occurs on the plant Xanthorrhoea glauca in natural ecosystems of Australia and is closely related to fusaria that produce a subgroup (type A) of trichothecene mycotoxins that lack a carbonyl group at carbon atom 8 (C-8). Here, mass spectrometric analysis reveale...

An old yellow enzyme gene controls the branch point between Aspergillus fumigatus and Claviceps purpurea ergot alkaloid pathways.

PubMed

Coyle, Christine M; Cheng, Johnathan Z; O'Connor, Sarah E; Panaccione, Daniel G

2010-06-01

Ergot fungi in the genus Claviceps and several related fungal groups in the family Clavicipitaceae produce toxic ergot alkaloids. These fungi produce a variety of ergot alkaloids, including clavines as well as lysergic acid derivatives. Ergot alkaloids are also produced by the distantly related, opportunistic human pathogen Aspergillus fumigatus. However, this fungus produces festuclavine and fumigaclavines A, B, and C, which collectively differ from clavines of clavicipitaceous fungi in saturation of the last assembled of four rings in the ergoline ring structure. The two lineages are hypothesized to share early steps of the ergot alkaloid pathway before diverging at some point after the synthesis of the tricyclic intermediate chanoclavine-I. Disruption of easA, a gene predicted to encode a flavin-dependent oxidoreductase of the old yellow enzyme class, in A. fumigatus led to accumulation of chanoclavine-I and chanoclavine-I-aldehyde. Complementation of the A. fumigatus easA mutant with a wild-type allele from the same fungus restored the wild-type profile of ergot alkaloids. These data demonstrate that the product of A. fumigatus easA is required for incorporation of chanoclavine-I-aldehyde into more-complex ergot alkaloids, presumably by reducing the double bond conjugated to the aldehyde group, thus facilitating ring closure. Augmentation of the A. fumigatus easA mutant with a homologue of easA from Claviceps purpurea resulted in accumulation of ergot alkaloids typical of clavicipitaceous fungi (agroclavine, setoclavine, and its diastereoisomer isosetoclavine). These data indicate that functional differences in the easA-encoded old yellow enzymes of A. fumigatus and C. purpurea result in divergence of their respective ergot alkaloid pathways.

An Old Yellow Enzyme Gene Controls the Branch Point between Aspergillus fumigatus and Claviceps purpurea Ergot Alkaloid Pathways▿

PubMed Central

Coyle, Christine M.; Cheng, Johnathan Z.; O'Connor, Sarah E.; Panaccione, Daniel G.

2010-01-01

Ergot fungi in the genus Claviceps and several related fungal groups in the family Clavicipitaceae produce toxic ergot alkaloids. These fungi produce a variety of ergot alkaloids, including clavines as well as lysergic acid derivatives. Ergot alkaloids are also produced by the distantly related, opportunistic human pathogen Aspergillus fumigatus. However, this fungus produces festuclavine and fumigaclavines A, B, and C, which collectively differ from clavines of clavicipitaceous fungi in saturation of the last assembled of four rings in the ergoline ring structure. The two lineages are hypothesized to share early steps of the ergot alkaloid pathway before diverging at some point after the synthesis of the tricyclic intermediate chanoclavine-I. Disruption of easA, a gene predicted to encode a flavin-dependent oxidoreductase of the old yellow enzyme class, in A. fumigatus led to accumulation of chanoclavine-I and chanoclavine-I-aldehyde. Complementation of the A. fumigatus easA mutant with a wild-type allele from the same fungus restored the wild-type profile of ergot alkaloids. These data demonstrate that the product of A. fumigatus easA is required for incorporation of chanoclavine-I-aldehyde into more-complex ergot alkaloids, presumably by reducing the double bond conjugated to the aldehyde group, thus facilitating ring closure. Augmentation of the A. fumigatus easA mutant with a homologue of easA from Claviceps purpurea resulted in accumulation of ergot alkaloids typical of clavicipitaceous fungi (agroclavine, setoclavine, and its diastereoisomer isosetoclavine). These data indicate that functional differences in the easA-encoded old yellow enzymes of A. fumigatus and C. purpurea result in divergence of their respective ergot alkaloid pathways. PMID:20435769

The effects of ergot and non-ergot-derived dopamine agonists in an experimental mouse model of endometriosis.

PubMed

Delgado-Rosas, Francisco; Gómez, Raúl; Ferrero, Hortensia; Gaytan, Francisco; Garcia-Velasco, Juan; Simón, Carlos; Pellicer, Antonio

2011-11-01

Implantation of a retrogradely shed endometrium during menstruation requires an adequate blood supply, which allows the growth of endometriotic lesions. This suggests that the development of endometriosis can be impaired by inhibiting angiogenesis. The growth of endometriotic foci is impaired by commercial oncological antiangiogenic drugs used to block vascular endothelial growth factor (VEGF) signaling. The dopamine agonist cabergoline (Cb2) inhibits the growth of established endometriosis lesions by exerting antiangiogenic effects through VEGFR2 inactivation. However, the use of ergot-derived Cb2 is associated with an increased incidence of cardiac valve regurgitation. To evaluate the potential usage of non-ergot-derived dopamine agonists for the treatment of human endometriosis, we compared the efficacy of quinagolide with that of Cb2 in preventing angiogenesis and vascularization in a heterologous mouse model of endometriosis. Nude mice whose peritoneum had been implanted with eutopic human endometrial fragments were treated with vehicle, 50  μg/kg per day oral Cb2, or 50 or 200  μg/kg per day quinagolide during a 14-day period. At the end of the treatment period, the implants were excised in order to assess lesion size, cell proliferation, degree of vascularization, and angiogenic gene expression. Neoangiogenesis was inhibited and the size of active endometriotic lesions, cellular proliferation index, and angiogenic gene expression were significantly reduced by both dopamine agonists when compared with the placebo. Given that Cb2 and quinagolide were equally effective in inhibiting angiogenesis and reducing lesion size, these experiments provide the rationale for pilot studies to explore the use of non-ergot-derived dopamine agonists for the treatment of endometriosis in humans.

Detection of new emerging type-A trichothecenes by untargeted mass spectrometry.

PubMed

González-Jartín, Jesús M; Alfonso, Amparo; Sainz, María J; Vieytes, Mercedes R; Botana, Luis M

2018-02-01

Mycotoxins occur naturally as agricultural contaminants all over the world. The toxic effects of some of their metabolites are known and their presence regulated in food and feed. This paper describes two methods for the detection of toxins of type-A trichothecenes group, and their modified forms, using mass spectrometry. Ultra-performance liquid chromatography coupled to mass spectrometry-ion trap-time of flight (UPLC-MS-IT-TOF) was employed to characterize the fragmentation pathways of 10 type-A trichothecenes, and characteristic ions were tentatively identified in scan mode through their accurate masses. Unknown signals were detected in a F. sporotrichioides extract, which afterwards were identified as seven modified forms of neosolaniol (NEO) and T-2 toxin. Then, UPLC coupled to tandem mass spectrometry (MS/MS) was employed to develop a precursor ion scanning method that can be used as a screening tool to detect any modified type-A trichothecenes. Copyright © 2017 Elsevier B.V. All rights reserved.

Relationships among Ergot Alkaloids, Cytochrome P450 Activity, and Beef Steer Growth

NASA Astrophysics Data System (ADS)

Rosenkrans, Charles; Ezell, Nicholas

2015-03-01

Determining a grazing animal’s susceptibility to ergot alkaloids has been a research topic for decades. Our objective was to determine if the Promega™ P450-Glo assay could be used to indirectly detect ergot alkaloids or their metabolites in urine of steers. The first experiment validated the effects of ergot alkaloids [0, 20, and 40 μM of ergotamine (ET), dihydroergotamine (DHET), and ergonovine (EN)] on human CYP3A4 using the P450-Glo assay (Promega™ V9800). With this assay, luminescence is directly proportional to CYP450 activity. Relative inhibition of in vitro cytochrome P450 activity was affected (P < 0.001) by an interaction between alkaloids and concentration. That interaction resulted in no concentration effect of EN, but within ET and DHET 20 and 40 µM concentrations inhibited CYP450 activity when compared with controls. In experiment 2, urine was collected from Angus-sired crossbred steers (n = 39; 216 ± 2.6 d of age; 203 ± 1.7 kg) after grazing tall fescue pastures for 105 d. Non-diluted urine was added to the Promega™ P450-Glo assay, and observed inhibition (3.7 % ± 2.7 of control). Urine content of total ergot alkaloids (331.1 ng/mg of creatinine ± 325.7) was determined using enzyme linked immunosorbent assay. Urine inhibition of CYP450 activity and total alkaloids were correlated (r = -0.31; P < 0.05). Steers were genotyped at CYP450 single nucleotide polymorphism, C994G. Steer genotype affected (P < 0.03) inhibition of CYP450 activity by urine; heterozygous steers had the least amount of CYP450 inhibition suggesting that genotyping cattle may be a method of identifying animals that are susceptible to ergot alkaloids. Although, additional research is needed, we demonstrate that the Promega™ P450-Glo assay is sensitive to ergot alkaloids and urine from steers grazing tall fescue. With some refinement the P450-Glo assay has potential as a tool for screening cattle for their exposure to fescue toxins.

Exposure to Ergot Alkaloids During Gestation Reduces Fetal Growth in Sheep

NASA Astrophysics Data System (ADS)

Duckett, Susan; Pratt, Scott; Andrae, John

2014-08-01

Tall fescue [Lolium arundinaceum (Schreb.) Darbysh; Schedonorus phoenix (Scop.) Holub] is the primary cool season perennial grass in the eastern U.S. Most tall fescue contains an endophyte (Neotyphodium coenophialum), which produces ergot alkaloids that cause vasoconstriction and could restrict blood flow to the fetus in pregnant animals. The objective of this study was to examine fetal growth during maternal exposure to ergot alkaloids during gestation. Pregnant ewes (n = 16) were randomly assigned to one of two dietary treatments: 1) endophyte-infected (Neotyphodium coenophialum) tall fescue seed (E+; 0.8 ug of ergovaline /g diet DM) and 2) endophyte-free tall fescue seed (E-; 0.0 ug of ergovaline/g diet DM). Birth weight of lambs was reduced by 37% for E+ compared to E-. Organ and muscle weights were also lighter for E+ than E-. Exposure to ergot alkaloids in utero reduces fetal growth and muscle development.

Effect of ergot alkaloids associated with fescue toxicosis on hepatic cytochrome P450 and antioxidant proteins

DOE Office of Scientific and Technical Information (OSTI.GOV)

Settivari, Raja S.; Evans, Tim J.; Rucker, Ed

Intake of ergot alkaloids found in endophyte-infected tall fescue grass is associated with decreased feed intake and reduction in body weight gain. The liver is one of the target organs of fescue toxicosis with upregulation of genes involved in xenobiotic metabolism and downregulation of genes associated with antioxidant pathways. It was hypothesized that short-term exposure of rats to ergot alkaloids would change hepatic cytochrome P450 (CYP) and antioxidant expression, as well as reduce antioxidant enzyme activity and hepatocellular proliferation rates. Hepatic gene expression of various CYPs, selected nuclear receptors associated with the CYP induction, and antioxidant enzymes were measured usingmore » real-time PCR. Hepatic expression of CYP, antioxidant and proliferating cell nuclear antigen (PCNA) proteins were measured using Western blots. The CYP3A1 protein expression was evaluated using primary rat hepatocellular cultures treated with ergovaline, one of the major ergot alkaloids produced by fescue endophyte, in order to assess the direct role of ergot alkaloids in CYP induction. The enzyme activities of selected antioxidants were assayed spectrophotometrically. While hepatic CYP and nuclear receptor expression were increased in ergot alkaloid-exposed rats, the expression and activity of antioxidant enzymes were reduced. This could potentially lead to increased oxidative stress, which might be responsible for the decrease in hepatocellular proliferation after ergot alkaloid exposure. This study demonstrated that even short-term exposure to ergot alkaloids can potentially induce hepatic oxidative stress which can contribute to the pathogenesis of fescue toxicosis.« less

Occurrence and distribution of 13 trichothecene toxins in naturally contaminated maize plants in Germany.

PubMed

Schollenberger, Margit; Müller, Hans-Martin; Ernst, Katrin; Sondermann, Sarah; Liebscher, Melanie; Schlecker, Claudia; Wischer, Gerald; Drochner, Winfried; Hartung, Karin; Piepho, Hans-Peter

2012-10-01

The objective of the present study was to monitor the occurrence and distribution of a spectrum of trichothecene toxins in different parts of maize plants. Therefore maize plants were sampled randomly from 13 fields in southwest Germany and the fractions kernels, cobs, husks, stalks, leaves and rudimentary ears were analyzed for eight A-type and five B-type trichothecenes. Each of the toxins was found in at least three of the total of 78 samples. The study revealed that both A-type and B-type trichothecenes may be present in all parts of the maize plant but may be unevenly distributed. For the contents of deoxynivalenol, 3- and 15-acetyldeoxynivalenol, nivalenol, scirpentriol, 15-monoacetoxyscirpenol, HT-2 and T-2 toxin significant differences (p < 0.05) were found between different parts of the maize plants whereas no significant differences were observed for fusarenon-X, 4,15-diacetoxyscirpenol, neosolaniol, T-2 triol and T-2 tetraol. Up to twelve toxins co-occurring in one sample were detected. As a group B-type trichothecenes dominated over A-type trichothecenes concerning incidences and levels. Contamination was strongest with rudimentary ears based on incidence and mean and maximum contents; mean contents with few exceptions tended towards a higher level than in other fractions with significant (p < 0.05) differences compared to leaves for seven toxins.

Effects of Ergot Alkaloids on Liver Function of Piglets as Evaluated by the 13C-Methacetin and 13C-α-Ketoisocaproic Acid Breath Test

PubMed Central

Dänicke, Sven; Diers, Sonja

2013-01-01

Ergot alkaloids (the sum of individual ergot alkaloids are termed as total alkaloids, TA) are produced by the fungus Claviceps purpurea, which infests cereal grains commonly used as feedstuffs. Ergot alkaloids potentially modulate microsomal and mitochondrial hepatic enzymes. Thus, the aim of the present experiment was to assess their effects on microsomal and mitochondrial liver function using the 13C-Methacetin (MC) and 13C-α-ketoisocaproic acid (KICA) breath test, respectively. Two ergot batches were mixed into piglet diets, resulting in 11 and 22 mg (Ergot 5-low and Ergot 5-high), 9 and 14 mg TA/kg (Ergot 15-low and Ergot 15-high) and compared to an ergot-free control group. Feed intake and live weight gain decreased significantly with the TA content (p < 0.001). Feeding the Ergot 5-high diet tended to decrease the 60-min-cumulative 13CO2 percentage of the dose recovery (cPDR60) by 26% and 28% in the MC and KICA breath test, respectively, compared to the control group (p = 0.065). Therefore, both microsomal and mitochondrial liver function was slightly affected by ergot alkaloids. PMID:23322130

Effects of ergot alkaloids on liver function of piglets as evaluated by the (13)C-methacetin and (13)C-α-ketoisocaproic acid breath test.

PubMed

Dänicke, Sven; Diers, Sonja

2013-01-15

Ergot alkaloids (the sum of individual ergot alkaloids are termed as total alkaloids, TA) are produced by the fungus Claviceps purpurea, which infests cereal grains commonly used as feedstuffs. Ergot alkaloids potentially modulate microsomal and mitochondrial hepatic enzymes. Thus, the aim of the present experiment was to assess their effects on microsomal and mitochondrial liver function using the (13)C-Methacetin (MC) and (13)C-α-ketoisocaproic acid (KICA) breath test, respectively. Two ergot batches were mixed into piglet diets, resulting in 11 and 22 mg (Ergot 5-low and Ergot 5-high), 9 and 14 mg TA/kg (Ergot 15-low and Ergot 15-high) and compared to an ergot-free control group. Feed intake and live weight gain decreased significantly with the TA content (p < 0.001). Feeding the Ergot 5-high diet tended to decrease the 60-min-cumulative (13)CO(2) percentage of the dose recovery (cPDR(60)) by 26% and 28% in the MC and KICA breath test, respectively, compared to the control group (p = 0.065). Therefore, both microsomal and mitochondrial liver function was slightly affected by ergot alkaloids.

Identification and in vitro cytotoxicity of ochratoxin A degradation products formed during coffee roasting.

PubMed

Cramer, Benedikt; Königs, Maika; Humpf, Hans-Ulrich

2008-07-23

The mycotoxin ochratoxin A is degraded by up to 90% during coffee roasting. In order to investigate this degradation, model heating experiments with ochratoxin A were carried out, and the reaction products were analyzed by HPLC-DAD and HPLC-MS/MS. Two ochratoxin A degradation products were identified, and their structure and absolute configuration were determined. As degradation reactions, the isomerization to 14-(R)-ochratoxin A and the decarboxylation to 14-decarboxy-ochratoxin A were identified. Subsequently, an analytical method for the determination of these compounds in roasted coffee was developed. Quantification was carried out by HPLC-MS/MS and the use of stable isotope dilution analysis. By using this method for the analysis of 15 coffee samples from the German market, it could be shown that, during coffee roasting, the ochratoxin A diastereomer 14-(R)-ochratoxin A was formed in amounts of up to 25.6% relative to ochratoxin A. The decarboxylation product was formed only in traces. For toxicity evaluations, first preliminary cell culture assays were performed with the two new substances. Both degradation products exhibited higher IC50 values and caused apoptotic effects with higher concentrations than ochratoxin A in cultured human kidney epithelial cells. Thus, these cell culture data suggest that the degradation products are less cytotoxic than ochratoxin A.

Effects of ergot alkaloids in feed on performance and liver function of piglets as evaluated by the ¹³C-methacetin breath test.

PubMed

Dänicke, Sven; Diers, Sonja

2013-02-01

Ergot alkaloids (the sum of individual alkaloids is termed as total alkaloids, TA) are mycotoxins of the fungus Claviceps purpurea and might adversely affect the performance and aspects of liver physiology of pigs. The objective of the study was to assess the effect of feeding ergot alkaloids to piglets on performance and liver function by using the ¹³C-methacetin breath test. Two ergot batches were mixed into piglet diets resulting in 5 and 6 mg (Ergot 17-low and -high) and 9 and 21 mg TA/kg (Ergot 19-low and -high) and compared to an ergot free Control group. Feed intake and live weight gain decreased significantly with the TA content (p = 0.006). The time of the maximum ¹³CO₂-exhalation (t (max)) occurred significantly earlier in Control piglets (8.9 min) compared to the groups Ergot 17-high and Ergot 19-high (24.7 and 23.6 min, respectively, p = 0.014) whilst the elimination half-life remained uninfluenced by dietary treatments (55-64 min). The cumulative ¹³CO₂-recovery (cPDR) was significantly reduced in piglets fed the Ergot 19-high diet (7.6%) compared to the groups Control and Ergot 17-high (13.1% and 10.8%, respectively, p = 0.011). In conclusion, the TA content of the diets is closer related to the adverse effects of ergot on piglet performance than the dietary ergot content itself. The mechanisms by which TA affects porcine liver function need to be studied further.

Effect of feeding sorghum ergot (Claviceps africana) to sows during mid-lactation on plasma prolactin and litter performance.

PubMed

Kopinski, J S; Blaney, B J; Murray, S-A; Downing, J A

2008-10-01

Diets containing 3% sorghum ergot (16 mg alkaloids/kg, including 14 mg dihydroergosine/kg) were fed to 12 sows from 14 days post-farrowing until weaning 14 days later, and their performance was compared with that of 10 control sows. Ergot-fed sows displayed a smaller weight loss during lactation of 24 kg/head vs. 29 kg/head in control sows (p > 0.05) despite feed consumption being less (61 kg/head total feed intake vs. 73 kg/head by control sows; p < 0.05). Ergot-fed sows had poorer weight gain of litters over the 14-day period (16.6 kg/litter vs. 28.3 kg/litter for controls; p < 0.05) despite an increase in consumption of creep feed by the piglets from the ergot-fed sows (1.9 kg/litter compared with 1.1 kg/litter by the control; p > 0.05). Sow plasma prolactin was reduced with ergot feeding after 7 days to 4.8 microg/l compared with 15.1 microg/l in the control sows (p < 0.01) and then at weaning was 4.9 microg/l compared with 8.0 microg/l (p < 0.01) in the control sows. Two sows fed ergot ceased lactation early, and the above sow feed intakes, body weight losses with litter weight gains and creep consumption indirectly indicate an ergot effect on milk production.

Permeability of ergot alkaloids across the blood-brain barrier in vitro and influence on the barrier integrity

PubMed Central

Mulac, Dennis; Hüwel, Sabine; Galla, Hans-Joachim; Humpf, Hans-Ulrich

2012-01-01

Scope Ergot alkaloids are secondary metabolites of Claviceps spp. and they have been in the focus of research for many years. Experiments focusing on ergotamine as a former migraine drug referring to the ability to reach the brain revealed controversial results. The question to which extent ergot alkaloids are able to cross the blood-brain barrier is still not answered. Methods and results In order to answer this question we have studied the ability of ergot alkaloids to penetrate the blood-brain barrier in a well established in vitro model system using primary porcine brain endothelial cells. It could clearly be demonstrated that ergot alkaloids are able to cross the blood-brain barrier in high quantities in only a few hours. We could further identify an active transport for ergometrine as a substrate for the BCRP/ABCG2 transporter. Investigations concerning barrier integrity properties have identified ergocristinine as a potent substance to accumulate in these cells ultimately leading to a weakened barrier function. Conclusion For the first time we could show that the so far as biologically inactive described 8-(S) isomers of ergot alkaloids seem to have an influence on barrier integrity underlining the necessity for a risk assessment of ergot alkaloids in food and feed. PMID:22147614

Activities and Effects of Ergot Alkaloids on Livestock Physiology and Production

PubMed Central

Klotz, James L.

2015-01-01

Consumption of feedstuffs contaminated with ergot alkaloids has a broad impact on many different physiological mechanisms that alters the homeostasis of livestock. This change in homeostasis causes an increased sensitivity in livestock to perturbations in the ambient environment, resulting in an increased sensitivity to such stressors. This ultimately results in large financial losses in the form of production losses to livestock producers around the world. This review will focus on the underlying physiological mechanisms that are affected by ergot alkaloids that lead to decreases in livestock production. PMID:26226000

D-lysergic acid-activating enzyme from the ergot fungus Claviceps purpurea.

PubMed Central

Keller, U; Zocher, R; Krengel, U; Kleinkauf, H

1984-01-01

A D-lysergic acid-activating enzyme from the ergot fungus Claviceps purpurea was purified about 145-fold. The enzyme was able to catalyse both the D-lysergic acid-dependent ATP-pyrophosphate exchange and the formation of ATP from D-lysergic acid adenylate and pyrophosphate. Both reactions were also catalysed to a decreased but significant extent with respect to dihydrolysergic acid. The molecular mass of the enzyme was estimated to lie between 135 and 140 kDa. The involvement of the enzyme in the biosynthesis of ergot peptide alkaloids is discussed. Images Fig. 4. PMID:6326747

Determinants and Expansion of Specificity in a Trichothecene UDP-Glucosyltransferase from Oryza sativa.

PubMed

Wetterhorn, Karl M; Gabardi, Kaitlyn; Michlmayr, Herbert; Malachova, Alexandra; Busman, Mark; McCormick, Susan P; Berthiller, Franz; Adam, Gerhard; Rayment, Ivan

2017-12-19

Family 1 UDP-glycosyltransferases (UGTs) in plants primarily form glucose conjugates of small molecules and, besides other functions, play a role in detoxification of xenobiotics. Indeed, overexpression of a barley UGT in wheat has been shown to control Fusarium head blight, which is a plant disease of global significance that leads to reduced crop yields and contamination with trichothecene mycotoxins such as deoxynivalenol (DON), T-2 toxin, and many other structural variants. The UGT Os79 from rice has emerged as a promising candidate for inactivation of mycotoxins because of its ability to glycosylate DON, nivalenol, and hydrolyzed T-2 toxin (HT-2). However, Os79 is unable to modify T-2 toxin (T-2), produced by pathogens such as Fusarium sporotrichioides and Fusarium langsethii. Activity toward T-2 is desirable because it would allow a single UGT to inactivate co-occurring mycotoxins. Here, the structure of Os79 in complex with the products UDP and deoxynivalenol 3-O-glucoside is reported together with a kinetic analysis of a broad range of trichothecene mycotoxins. Residues associated with the trichothecene binding pocket were examined by site-directed mutagenesis that revealed that trichothecenes substituted at the C4 position, which are not glycosylated by wild-type Os79, can be accommodated in the binding pocket by increasing its volume. The H122A/L123A/Q202L triple mutation, which increases the volume of the active site and attenuates polar contacts, led to strong and equivalent activity toward trichothecenes with C4 acetyl groups. This mutant enzyme provides the broad specificity required to control multiple toxins produced by different Fusarium species and chemotypes.

Detection of airborne Stachybotrys chartarum macrocyclic trichothecene mycotoxins on particulates smaller than conidia.

PubMed

Brasel, T L; Douglas, D R; Wilson, S C; Straus, D C

2005-01-01

Highly respirable particles (diameter, <1 microm) constitute the majority of particulate matter found in indoor air. It is hypothesized that these particles serve as carriers for toxic compounds, specifically the compounds produced by molds in water-damaged buildings. The presence of airborne Stachybotrys chartarum trichothecene mycotoxins on particles smaller than conidia (e.g., fungal fragments) was therefore investigated. Cellulose ceiling tiles with confluent Stachybotrys growth were placed in gas-drying containers through which filtered air was passed. Exiting particulates were collected by using a series of polycarbonate membrane filters with decreasing pore sizes. Scanning electron microscopy was employed to determine the presence of conidia on the filters. A competitive enzyme-linked immunosorbent assay (ELISA) specific for macrocyclic trichothecenes was used to analyze filter extracts. Cross-reactivity to various mycotoxins was examined to confirm the specificity. Statistically significant (P < 0.05) ELISA binding was observed primarily for macrocyclic trichothecenes at concentrations of 50 and 5 ng/ml and 500 pg/ml (58.4 to 83.5% inhibition). Of the remaining toxins tested, only verrucarol and diacetylverrucarol (nonmacrocyclic trichothecenes) demonstrated significant binding (18.2 and 51.7% inhibition, respectively) and then only at high concentrations. The results showed that extracts from conidium-free filters demonstrated statistically significant (P < 0.05) antibody binding that increased with sampling time (38.4 to 71.9% inhibition, representing a range of 0.5 to 4.0 ng/ml). High-performance liquid chromatography analysis suggested the presence of satratoxin H in conidium-free filter extracts. These data show that S. chartarum trichothecene mycotoxins can become airborne in association with intact conidia or smaller particles. These findings may have important implications for indoor air quality assessment.

Engaging One Health for Non-Communicable Diseases in Africa: Perspective for Mycotoxins

PubMed Central

Ladeira, Carina; Frazzoli, Chiara; Orisakwe, Orish Ebere

2017-01-01

The role of mycotoxins—e.g., aflatoxins, ochratoxins, trichothecenes, zearalenone, fumonisins, tremorgenic toxins, and ergot alkaloids—has been recognized in the etiology of a number of diseases. In many African countries, the public health impact of chronic (indoor) and/or repeated (dietary) mycotoxin exposure is largely ignored hitherto, with impact on human health, food security, and export of African agricultural food products. Notwithstanding, African scientific research reached milestones that, when linked to findings gained by the international scientific community, make the design and implementation of science-driven governance schemes feasible. Starting from Nigeria as leading African Country, this article (i) overviews available data on mycotoxins exposure in Africa; (ii) discusses new food safety issues, such as the environment–feed–food chain and toxic exposures of food producing animals in risk assessment and management; (iii) identifies milestones for mycotoxins risk management already reached in West Africa; and (iv) points out preliminary operationalization aspects for shielding communities from direct (on health) and indirect (on trade, economies, and livelihoods) effects of mycotoxins. An African science-driven engaging of scientific knowledge by development actors is expected therefore. In particular, One health/One prevention is suggested, as it proved to be a strategic and sustainable development framework. PMID:29085817

Improving field production of ergot alkaloids by application of gametocide on rye host plants.

PubMed

Hanosová, Helena; Koprna, Radoslav; Valík, Josef; Knoppová, Lucie; Frébort, Ivo; Dzurová, Lenka; Galuszka, Petr

2015-12-25

Ergot alkaloids are widely used in the pharmaceutical industry in drug preparations for treating migraines and Parkinson's disease, inducing uterine contraction, and other purposes. Phytopathogenic fungi of the genus Claviceps (e.g. C. purpurea) comprise a major biological source of ergot alkaloids. Worldwide industrial production of these alkaloids derives almost equally from two biotechnological procedures: submerged culture of the fungus in fermenters and field parasitic production in dormant fungal organs known as sclerotia (also termed ergot). Ergot yields from field cultivation are greatly affected by weather and also can be much reduced by pollen contamination from imperfectly male-sterile rye, as only unfertilized ovaries can be infected by C. purpurea spores. Two substances with gametocidal effect - maleic hydrazide and 2-chloroethylphosphonic acid - were tested during three consecutive seasons in small field experiments for the ability to induce or amplify the male sterility of rye as well as the impacts on germination of C. purpurea spores and general vitality of rye host plants. Maleic hydrazide was proven to be a highly effective gametocide on both a fertile rye variety and a variety with imperfectly induced cytoplasmic male sterility. It showed negligible effect on germination of C. purpurea spores. Both accurate dosaging of the active gametocidal compound and timing of the application just 2-3 weeks before onset of anthesis proved crucial to achieving high ergot yield with minimum grain impurities. Copyright © 2015 Elsevier B.V. All rights reserved.

Ergot alkaloids decrease rumen epithelial blood flow

USDA-ARS?s Scientific Manuscript database

Two experiments were conducted to determine if ergot alkaloids affect blood flow to the absorptive surface of the rumen of steers. Steers (n=8 total) were pair-fed alfalfa cubes at 1.5× NEM and received ground endophyte-infected tall fescue seed (E+) or endophyte-free tall fescue seed (E-) via rumen...

Effect of Ergot Alkaloids on Bovine Foregut Vasculature

USDA-ARS?s Scientific Manuscript database

Ergot alkaloids induce vasoconstriction of bovine foregut vasculature. Ergovaline induced the greatest response in ruminal artery while ergovaline and ergotamine induced the greatest response in ruminal vein. Lysergic acid did not stimulate a contractile response in either the ruminal artery or vein...

Semiquantitative determination of ergot alkaloids in seed, straw, and digesta samples using a competitive enzyme-linked immunosorbent assay.

PubMed

Schnitzius, J M; Hill, N S; Thompson, C S; Craig, A M

2001-05-01

Ergot alkaloids present in endophyte-infected (E+) tall fescue cause fescue toxicosis and other toxic effects in livestock that consume infected plant tissue, leading to significant financial losses in livestock production each year. The predominant method currently in use for quantifying ergot alkaloid content in plant tissue is through high-performance liquid chromatography (HPLC), which quantifies the amount of ergovaline, one of many ergot alkaloids in E+ plant tissue. The enzyme-linked immunosorbent assay (ELISA) method used in this study detects quantities of nonspecific ergot alkaloids and therefore accounts for greater amounts of the total ergot alkaloid content in E+ tissue than does HPLC. The ELISA can also be used to more expediently analyze a larger number of forage samples without sophisticated and costly analytical equipment and therefore could be more desirable in a diagnostic setting. The purpose of this study was to evaluate the between-day and within-run variability of the ELISA and to determine the binding efficiency of 6 ergot alkaloids to the 15F3.E5 antibody used in the competitive ELISA to ascertain its feasibility as a quick analysis tool for ergot alkaloids. Straw samples had an average coefficient of variation (CV) for concentration of 10.2% within runs and 18.4% between runs, and the seed samples had an average CV for concentration of 13.3% within runs and 24.5% between runs. The grass tissue-based lysergic acid standard curve calculated from the ELISA had an average r2 of 0.99, with a CV of 2.1%. Ergocryptine, ergocristine, ergocornine, and ergotamine tartrate did not bind strongly to the 15F3.E5 antibody because of the presence of large side groups on these molecules, which block their binding to the antibody, whereas ergonovine and ergonovine maleate were bound much more efficiently because of their structural similarity to lysergic acid. Clarified rumen fluid was tested as an additional matrix for use in the ergot alkaloid competitive

Population genetic analysis and trichothecene profiling of Fusarium graminearum from wheat in Uruguay.

PubMed

Pan, D; Mionetto, A; Calero, N; Reynoso, M M; Torres, A; Bettucci, L

2016-03-11

Fusarium graminearum sensu stricto (F. graminearum s.s.) is the major causal agent of Fusarium head blight of wheat worldwide, and contaminates grains with trichothecene mycotoxins that cause serious threats to food safety and animal health. An important aspect of managing this pathogen and reducing mycotoxin contamination of wheat is knowledge regarding its population genetics. Therefore, isolates of F. graminearum s.s. from the major wheat-growing region of Uruguay were analyzed by amplified fragment length polymorphism assays, PCR genotyping, and chemical analysis of trichothecene production. Of the 102 isolates identified as having the 15-ADON genotype via PCR genotyping, all were DON producers, but only 41 strains were also 15-ADON producers, as determined by chemical analysis. The populations were genotypically diverse but genetically similar, with significant genetic exchange occurring between them. Analysis of molecular variance indicated that most of the genetic variability resulted from differences between isolates within populations. Multilocus linkage disequilibrium analysis suggested that the isolates had a panmictic population genetic structure and that there is significant recombination occurs in F. graminearum s.s. In conclusion, tour findings provide the first detailed description of the genetic structure and trichothecene production of populations of F. graminearum s.s. from Uruguay, and expands our understanding of the agroecology of F. graminearum and of the correlation between genotypes and trichothecene chemotypes.

Fate of mycotoxins during beer brewing and fermentation.

PubMed

Inoue, Tomonori; Nagatomi, Yasushi; Uyama, Atsuo; Mochizuki, Naoki

2013-01-01

Mycotoxins are frequent contaminants of grains, and breweries need, therefore, to pay close attention to the risk of contamination in beer made from such grains as barley and corn. The fate of 14 types of mycotoxin (aflatoxins, fumonisins, ochratoxin A, patulin, trichothecenes, and zearalenone) during beer brewing was investigated in this study. Malt artificially spiked with each mycotoxin was put through the mashing, filtration, boiling and fermentation processes involved in brewing. After brewing, the levels of aflatoxins, ochratoxin A, patulin, and zearalenone were found to have decreased to less than 20% of their initial concentration. They had been adsorbed mainly to the spent grain and removed from the unhopped wort. Additionally, as zearalenone was known, patulin was metabolized to the less toxic compound during the fermentation process. The risk of carry-over to beer was therefore reduced for half of the mycotoxins studied. However, attention still needs to be paid to the risk of trichothecene contamination.

Ochratoxin degradation and adsorption caused by astaxanthin-producing yeasts.

PubMed

Péteri, Z; Téren, J; Vágvölgyi, C; Varga, J

2007-05-01

Ochratoxin degrading and adsorbing activities of Phaffia rhodozyma and Xanthophyllomyces dendrorhous isolates were tested. P. rhodozyma CBS 5905 degraded more than 90% of ochratoxin A (OTA) in 15 days at 20 degrees C. The data presented indicate that P. rhodozyma is able to convert OTA to ochratoxin alpha, and this conversion is possibly mediated by an enzyme related to carboxypeptidases. Chelating agents like EDTA and 1,10-phenanthroline inhibited OTA degradation caused by P. rhodozyma indicating that the carboxypeptidase is a metalloprotease, similarly to carboxypeptidase A. The temperature optimum of this enzyme was found to be above 30 degrees C, which is much higher than the temperature optimum for growth of P. rhodozyma cells, which is around 20 degrees C. The enzyme responsible for ochratoxin degradation was found to be cell-bound. Besides, both viable and heat-treated (dead) P. rhodozyma cells were also able to adsorb significant amounts (up to 250 ng ml(-1)) of OTA. Heat treatment enhanced OTA adsorbing activities of the cells. Further studies are in progress to identify the enzyme responsible for OTA degradation in P. rhodozyma.

Ochratoxin A Detection on Antibody- Immobilized on BSA-Functionalized Gold Electrodes.

PubMed

Badea, Mihaela; Floroian, Laura; Restani, Patrizia; Cobzac, Simona Codruta Aurora; Moga, Marius

2016-01-01

Ochratoxin A (OTA)-a toxin produced by Aspergillus carbonarius, Aspergillus ochraceus, and Penicillium verrucosum-is one of the most-abundant food-contaminating mycotoxins. To avoid the risk of OTA consumption for humans and animals, the rapid detection and quantitation of OTA level in different commodities are of great importance. In this work, an impedimetric immunosensor for ochratoxin A (OTA) detection, a common toxic botanical contaminant, was developed via the immobilization of anti-OTA antibody on bovine serum albumin modified gold electrodes. A four-step reaction protocol was tested to modify the gold electrode and obtain the sensing substrate. All the steps of the immunosensor elaboration and also the immunochemical reaction between surface-bound antibody and ochratoxin A were analyzed using cyclic voltammetry and electrochemical impedance spectroscopy. Modification of the impedance due to the specific antigen-antibody reaction at immunosensor surface, was used in order to detect ochratoxin A. Linear proportionality of the charge transfer resistance to the concentration of OTA allows ochratoxin A detection in the range of 2.5-100 ng/mL.

Identification of cellular and molecular factors determining the response of cancer cells to six ergot alkaloids.

PubMed

Mrusek, Marco; Seo, Ean-Jeong; Greten, Henry Johannes; Simon, Michael; Efferth, Thomas

2015-02-01

Ergot alkaloids are psychoactive and vasoconstricting agents of the fungus Claviceps purpurea causing poisoning such as ergotism in medieval times (St. Anthony's Fire). This class of substances also inhibits tumor growth in vitro and in vivo, though the underlying mechanisms are unclear as yet. We investigated six ergot alkaloids (agroclavine, ergosterol, ergocornin E, ergotamine, dihydroergocristine, and 1-propylagroclavine tartrate) for their cytotoxicity towards tumor cell lines of the National Cancer Institute, USA. 1-Propylagroclavine tartrate (1-PAT) revealed the strongest cytotoxicity. Out of 76 clinically established anticancer drugs, cross-resistance was found between the ergot alkaloids and 6/7 anti-hormonal drugs (=85.7 %) and 5/15 DNA-alkylating drugs (=33.3 %). The IC50 values for the six alkaloids were not correlated to well-known determinants of drug resistance, such as proliferative activity (as measured by cell doubling times, PCNA expression, and cell cycle distribution), the multidrug resistance-mediating P-glycoprotein/MDR1 and expression or mutations of oncogenes and tumor suppressor genes (EGFR, RAS, TP53). While resistance of control drugs (daunorubicin, cisplatin, erlotinib) correlated with these classical resistance mechanisms, ergot alkaloids did not. Furthermore, COMPARE and hierarchical cluster analyses were performed of mRNA microarray data to identify genes correlating with sensitivity or resistance to 1-PAT. Twenty-three genes were found with different biological functions (signal transducers, RNA metabolism, ribosome constituents, cell cycle and apoptosis regulators etc.). The expression of only 3/66 neuroreceptor genes correlated with the IC50 values for 1-PAT, suggesting that the psychoactive effects of ergot alkaloids may not play a major role for the cytotoxic activity against cancer cells. In conclusion, the cytotoxicity of ergot alkaloids is not involved in classical mechanisms of drug resistance opening the possibility to

Structural and functional characterization of TRI3 trichothecene 15-O-acetyltransferase from Fusarium sporotrichioides

DOE Office of Scientific and Technical Information (OSTI.GOV)

Garvey, Graeme S.; McCormick, Susan P.; Alexander, Nancy J.

2009-08-14

Fusarium head blight is a devastating disease of cereal crops whose worldwide incidence is increasing and at present there is no satisfactory way of combating this pathogen or its associated toxins. There is a wide variety of trichothecene mycotoxins and they all contain a 12,13-epoxytrichothecene skeleton but differ in their substitutions. Indeed, there is considerable variation in the toxin profile across the numerous Fusarium species that has been ascribed to differences in the presence or absence of biosynthetic enzymes and their relative activity. This article addresses the source of differences in acetylation at the C15 position of the trichothecene molecule.more » Here, we present the in vitro structural and biochemical characterization of TRI3, a 15-O-trichothecene acetyltransferase isolated from F. sporotrichioides and the 'in vivo' characterization of Deltatri3 mutants of deoxynivalenol (DON) producing F. graminearum strains. A kinetic analysis shows that TRI3 is an efficient enzyme with the native substrate, 15-decalonectrin, but is inactive with either DON or nivalenol. The structure of TRI3 complexed with 15-decalonectrin provides an explanation for this specificity and shows that Tri3 and Tri101 (3-O-trichothecene acetyltransferase) are evolutionarily related. The active site residues are conserved across all sequences for TRI3 orthologs, suggesting that differences in acetylation at C15 are not due to differences in Tri3. The tri3 deletion mutant shows that acetylation at C15 is required for DON biosynthesis even though DON lacks a C15 acetyl group. The enzyme(s) responsible for deacetylation at the 15 position of the trichothecene mycotoxins have not been identified.« less

Ochratoxin A production by Penicillium thymicola.

PubMed

Nguyen, Hai D T; McMullin, David R; Ponomareva, Ekaterina; Riley, Robert; Pomraning, Kyle R; Baker, Scott E; Seifert, Keith A

2016-08-01

Ochratoxin A (OTA) is a mycotoxin produced by some Aspergillus and Penicillium species that grow on economically important agricultural crops and food products. OTA is classified as Group 2B carcinogen and is potently nephrotoxic, which is the basis for its regulation in some jurisdictions. Using high resolution mass spectroscopy, OTA and ochratoxin B (OTB) were detected in liquid culture extracts of Penicillium thymicola DAOMC 180753 isolated from Canadian cheddar cheese. The genome of this strain was sequenced, assembled and annotated to probe for putative genes involved in OTA biosynthesis. Known OTA biosynthetic genes from Penicillium verrucosum or Penicillium nordicum, two related Penicillium species that produce OTA, were not found in P. thymicola. However, a gene cluster containing a polyketide synthase (PKS) and PKS-nonribosomal peptide synthase (NRPS) hybrid encoding genes were located in the P. thymicola genome that showed a high degree of similarity to OTA biosynthetic enzymes of Aspergillus carbonarius and Aspergillus ochraceus. This is the first report of ochratoxin from P. thymicola and a new record of the species in Canada. Crown Copyright © 2016. Published by Elsevier Ltd. All rights reserved.

Quantification of Trichothecene-Producing Fusarium Species in Harvested Grain by Competitive PCR To Determine Efficacies of Fungicides against Fusarium Head Blight of Winter Wheat

PubMed Central

Edwards, S. G.; Pirgozliev, S. R.; Hare, M. C.; Jenkinson, P.

2001-01-01

We developed a PCR-based assay to quantify trichothecene-producing Fusarium based on primers derived from the trichodiene synthase gene (Tri5). The primers were tested against a range of fusarium head blight (FHB) (also known as scab) pathogens and found to amplify specifically a 260-bp product from 25 isolates belonging to six trichothecene-producing Fusarium species. Amounts of the trichothecene-producing Fusarium and the trichothecene mycotoxin deoxynivalenol (DON) in harvested grain from a field trial designed to test the efficacies of the fungicides metconazole, azoxystrobin, and tebuconazole to control FHB were quantified. No correlation was found between FHB severity and DON in harvested grain, but a good correlation existed between the amount of trichothecene-producing Fusarium and DON present within grain. Azoxystrobin did not affect levels of trichothecene-producing Fusarium compared with those of untreated controls. Metconazole and tebuconazole significantly reduced the amount of trichothecene-producing Fusarium in harvested grain. We hypothesize that the fungicides affected the relationship between FHB severity and the amount of DON in harvested grain by altering the proportion of trichothecene-producing Fusarium within the FHB disease complex and not by altering the rate of DON production. The Tri5 quantitative PCR assay will aid research directed towards reducing amounts of trichothecene mycotoxins in food and animal feed. PMID:11282607

Label-free impedimetric immunosensor for sensitive detection of ochratoxin A.

PubMed

Radi, Abd-Elgawad; Muñoz-Berbel, Xavier; Lates, Vasilica; Marty, Jean-Louis

2009-03-15

A novel label-free electrochemical impedimetric immunosensor for sensitive detection of ochratoxin A (OTA) was reported. A two-step reaction protocol was elaborated to modify the gold electrode. The electrode was first derivatized by electrochemical reduction of in situ generated 4-carboxyphenyl diazonium salt (4-CPDS) in acidic aqueous solution yielded stable 4-carboxyphenyl (4-CP) monolayer. The ochratoxin A antibody was then immobilized making use of the carbodiimide chemistry. The steps of the immunosensor elaboration and the immunochemical reaction between ochratoxin A and the surface-bound antibody were interrogated using cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). The impedance change, due to the specific immuno-interaction at the immunosensor surface was utilized to detect ochratoxin A. The increase in electron-transfer resistance (DeltaR(et)) values was linearly proportional to the concentration of OTA in the range of 1-20ngmL(-1), with a detection limit of 0.5ngmL(-1).

Vasoconstrictive responses of the testicular and caudal arteries in bulls exposed to ergot alkaloids from tall fescue

USDA-ARS?s Scientific Manuscript database

Color Doppler ultasonography was used to evaluate vasoconstrictive responses of the testicular artery in yearling bulls to ergot alkaloids. Ergot alkaloid-induced constriction of the testicular artery could disrupt thermoregulation of the testes and reduce bull fertility. Luminal areas of the test...

Ergotism related to a single dose of ergotamine tartrate in an AIDS patient treated with ritonavir

PubMed Central

Blanche, P; Rigolet, A; Gombert, B; Ginsburg, C; Salmon, D; Sicard, D

1999-01-01

We report a rare case of ergotism related to a single dose of ergotamine tartrate in a man with AIDS being treated with ritonavir. He was treated with a prostacyclin analogue and made a complete recovery.


Keywords: ergotism; ergotamine tartrate; AIDS; ritonavir; adverse drug reaction; HIV infection PMID:10616689

Identification of loci and functional characterization of trichothecene biosynthesis genes in the filamentous fungus of the genus Trichoderma

USDA-ARS?s Scientific Manuscript database

Trichothecenes are mycotoxins produced by Trichoderma, Fusarium and at least four other genera in the fungal order Hypocreales. Fusarium has a trichothecene biosynthetic gene (TRI) cluster that encodes transport and regulatory proteins as well as most enzymes required for formation of the mycotoxin...

Identification of Loci and Functional Characterization of Trichothecene Biosynthesis Genes in Filamentous Fungi of the Genus Trichoderma▿†

PubMed Central

Cardoza, R. E.; Malmierca, M. G.; Hermosa, M. R.; Alexander, N. J.; McCormick, S. P.; Proctor, R. H.; Tijerino, A. M.; Rumbero, A.; Monte, E.; Gutiérrez, S.

2011-01-01

Trichothecenes are mycotoxins produced by Trichoderma, Fusarium, and at least four other genera in the fungal order Hypocreales. Fusarium has a trichothecene biosynthetic gene (TRI) cluster that encodes transport and regulatory proteins as well as most enzymes required for the formation of the mycotoxins. However, little is known about trichothecene biosynthesis in the other genera. Here, we identify and characterize TRI gene orthologues (tri) in Trichoderma arundinaceum and Trichoderma brevicompactum. Our results indicate that both Trichoderma species have a tri cluster that consists of orthologues of seven genes present in the Fusarium TRI cluster. Organization of genes in the cluster is the same in the two Trichoderma species but differs from the organization in Fusarium. Sequence and functional analysis revealed that the gene (tri5) responsible for the first committed step in trichothecene biosynthesis is located outside the cluster in both Trichoderma species rather than inside the cluster as it is in Fusarium. Heterologous expression analysis revealed that two T. arundinaceum cluster genes (tri4 and tri11) differ in function from their Fusarium orthologues. The Tatri4-encoded enzyme catalyzes only three of the four oxygenation reactions catalyzed by the orthologous enzyme in Fusarium. The Tatri11-encoded enzyme catalyzes a completely different reaction (trichothecene C-4 hydroxylation) than the Fusarium orthologue (trichothecene C-15 hydroxylation). The results of this study indicate that although some characteristics of the tri/TRI cluster have been conserved during evolution of Trichoderma and Fusarium, the cluster has undergone marked changes, including gene loss and/or gain, gene rearrangement, and divergence of gene function. PMID:21642405

Effects of acquisition, loss, and neofunctionalization of trichothecene biosynthetic genes on variation in trichothecene structure, pathway regulation, and self-protection mechanisms in the Hypocreales

USDA-ARS?s Scientific Manuscript database

Trichothecenes are secondary metabolites produced by multiple genera in the order Hypocreales, including Fusarium, Myrothecium, Stachybotrys, and Trichoderma. These metabolites are of concern because they are toxic to humans and animals, can contribute to pathogenicity in Fusarium, and are required ...

Occurrence of ochratoxin A in cocoa by-products and determination of its reduction during chocolate manufacture.

PubMed

Copetti, Marina V; Iamanaka, Beatriz T; Nester, Melanie A; Efraim, Priscilla; Taniwaki, Marta H

2013-01-01

This work reports an investigation carried out to assess the natural occurrence of ochratoxin A in 168 samples from different fractions obtained during the technological processing of cocoa (shell, nibs, liquor, butter, cake and cocoa powder) and the reduction of ochratoxin A during chocolate manufacture. Ochratoxin A analyses were performed with immunoaffinity columns and detection by high performance liquid chromatography. Concerning the natural ochratoxin A contamination in cocoa by-products, the highest levels of ochratoxin A were found in the shell, cocoa powder and cocoa cake. The cocoa butter was the least contaminated, showing that ochratoxin A seems to remain in the defatted cocoa solids. Under the technological conditions applied during the manufacture of chocolate in this study and the level of contamination present in the cocoa beans, this experiment demonstrated that 93.6% of ochratoxin A present in the beans was reduced during the chocolate producing. Copyright © 2012 Elsevier Ltd. All rights reserved.

Localization of ergot alkaloids in sclerotia of Claviceps purpurea by matrix-assisted laser desorption/ionization mass spectrometry imaging.

PubMed

Dopstadt, Julian; Vens-Cappell, Simeon; Neubauer, Lisa; Tudzynski, Paul; Cramer, Benedikt; Dreisewerd, Klaus; Humpf, Hans-Ulrich

2017-02-01

The fungus Claviceps purpurea produces highly toxic ergot alkaloids and accumulates these in the hardened bodies of fungal mycelium. These so-called sclerotia, or ergot bodies, replace the crop seed of infected plants, which can include numerous important food- and feedstuff such as rye and wheat. While several studies have explored details of the infection process and development of ergot bodies, little information is available on the spatial distribution of the mycotoxins in the sclerotia. Here we used matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) at a lateral resolution of 35 μm to visualize the distribution of two representative alkaloids, ergocristine and ergometrine, produced by Ecc93 and Gal 310 variants of C. purpurea, respectively, after infection of rye. To improve cryosectioning of this fragile biological material tissue with complex texture, we developed a practical embedding protocol based on cellulose polymers. The MALDI-MS images recorded from the so produced intact tissues sections revealed that ergometrine exhibited a relatively homogeneous distribution throughout the ergot body, whereas ergocristine was found to be enriched in the proximal region. This finding can be correlated to the morphological development of sclerotia as ergot alkaloids are only produced in the sphacelial stage. The ability to localize toxins and other secondary metabolites in intact sections of crop-infecting fungi with high lateral resolution renders MALDI-MSI a powerful tool for investigating biosynthetic pathways and for obtaining a deeper understanding of the parasite-host interaction. Graphical abstract Workflow for identification and spatial localization of ergot alkaloids in infected rye grains.

Ergot alkaloid transport across ruminant gastric tissues.

PubMed

Hill, N S; Thompson, F N; Stuedemann, J A; Rottinghaus, G W; Ju, H J; Dawe, D L; Hiatt, E E

2001-02-01

Ergot alkaloids cause fescue toxicosis when livestock graze endophyte-infected tall fescue. It is generally accepted that ergovaline is the toxic component of endophyte-infected tall fescue, but there is no direct evidence to support this hypothesis. The objective of this study was to examine relative and potential transport of ergoline and ergopeptine alkaloids across isolated gastric tissues in vitro. Sheep ruminal and omasal tissues were surgically removed and placed in parabiotic chambers. Equimolar concentrations of lysergic acid, lysergol, ergonovine, ergotamine, and ergocryptine were added to a Kreb's Ringer phosphate (KRP) solution on the mucosal side of the tissue. Tissue was incubated in near-physiological conditions for 240 min. Samples were taken from KRP on the serosal side of the chambers at times 0, 30, 60, 120, 180, and 240 min and analyzed for ergot alkaloids by competitive ELISA. The serosal KRP remaining after incubation was freeze-dried and the alkaloid species quantified by HPLC. The area of ruminal and omasal tissues was measured and the potential transportable alkaloids calculated by multiplying the moles of transported alkaloids per square centimeter of each tissue type by the surface area of the tissue. Studies were conducted to compare alkaloid transport in reticular, ruminal, and omasal tissues and to determine whether transport was active or passive. Ruminal tissue had greater ergot alkaloid transport potential than omasal tissue (85 vs 60 mmol) because of a larger surface area. The ruminal posterior dorsal sac had the greatest potential for alkaloid transport, but the other ruminal tissues were not different from one another. Alkaloid transport was less among reticular tissues than among ruminal tissues. Transport of alkaloids seemed to be an active process. The alkaloids with greatest transport potential were lysergic acid and lysergol. Ergopeptine alkaloids tended to pass across omasal tissues in greater quantities than across ruminal

Molecularly Imprinted Polymers for Ochratoxin A Extraction and Analysis

PubMed Central

Yu, Jorn C. C.; Lai, Edward P. C.

2010-01-01

Molecularly imprinted polymers (MIPs) are considered as polymeric materials that mimic the functionality of antibodies. MIPs have been utilized for a wide variety of applications in chromatography, solid phase extraction, immunoassays, and sensor recognition. In this article, recent advances of MIPs for the extraction and analysis of ochratoxins are discussed. Selection of functional monomers to bind ochratoxin A (OTA) with high affinities, optimization of extraction procedures, and limitations of MIPs are compared from different reports. The most relevant examples in the literature are described to clearly show how useful these materials are. Strategies on MIP preparation and schemes of analytical methods are also reviewed in order to suggest the next step that would make better use of MIPs in the field of ochratoxin research. The review ends by outlining the remaining issues and impediments. PMID:22069649

UDP-Glucosyltransferases from Rice, Brachypodium, and Barley: Substrate Specificities and Synthesis of Type A and B Trichothecene-3-O-β-d-glucosides

PubMed Central

Malachová, Alexandra; Piątkowska, Marta; Hametner, Christian; Šofrová, Jana; Jaunecker, Günther; Häubl, Georg; Lemmens, Marc

2018-01-01

Trichothecene toxins are confirmed or suspected virulence factors of various plant-pathogenic Fusarium species. Plants can detoxify these to a variable extent by glucosylation, a reaction catalyzed by UDP-glucosyltransferases (UGTs). Due to the unavailability of analytical standards for many trichothecene-glucoconjugates, information on such compounds is limited. Here, the previously identified deoxynivalenol-conjugating UGTs HvUGT13248 (barley), OsUGT79 (rice) and Bradi5g03300 (Brachypodium), were expressed in E. coli, affinity purified, and characterized towards their abilities to glucosylate the most relevant type A and B trichothecenes. HvUGT13248, which prefers nivalenol over deoxynivalenol, is also able to conjugate C-4 acetylated trichothecenes (e.g., T-2 toxin) to some degree while OsUGT79 and Bradi5g03300 are completely inactive with C-4 acetylated derivatives. The type A trichothecenes HT-2 toxin and T-2 triol are the kinetically preferred substrates in the case of HvUGT13248 and Bradi5g03300. We glucosylated several trichothecenes with OsUGT79 (HT-2 toxin, T-2 triol) and HvUGT13248 (T-2 toxin, neosolaniol, 4,15-diacetoxyscirpenol, fusarenon X) in the preparative scale. NMR analysis of the purified glucosides showed that exclusively β-d-glucosides were formed regio-selectively at position C-3-OH of the trichothecenes. These synthesized standards can be used to investigate the occurrence and toxicological properties of these modified mycotoxins. PMID:29509722

UDP-Glucosyltransferases from Rice, Brachypodium, and Barley: Substrate Specificities and Synthesis of Type A and B Trichothecene-3-O-β-d-glucosides.

PubMed

Michlmayr, Herbert; Varga, Elisabeth; Malachová, Alexandra; Fruhmann, Philipp; Piątkowska, Marta; Hametner, Christian; Šofrová, Jana; Jaunecker, Günther; Häubl, Georg; Lemmens, Marc; Berthiller, Franz; Adam, Gerhard

2018-03-06

Trichothecene toxins are confirmed or suspected virulence factors of various plant-pathogenic Fusarium species. Plants can detoxify these to a variable extent by glucosylation, a reaction catalyzed by UDP-glucosyltransferases (UGTs). Due to the unavailability of analytical standards for many trichothecene-glucoconjugates, information on such compounds is limited. Here, the previously identified deoxynivalenol-conjugating UGTs HvUGT13248 (barley), OsUGT79 (rice) and Bradi5g03300 ( Brachypodium ), were expressed in E. coli , affinity purified, and characterized towards their abilities to glucosylate the most relevant type A and B trichothecenes. HvUGT13248, which prefers nivalenol over deoxynivalenol, is also able to conjugate C-4 acetylated trichothecenes (e.g., T-2 toxin) to some degree while OsUGT79 and Bradi5g03300 are completely inactive with C-4 acetylated derivatives. The type A trichothecenes HT-2 toxin and T-2 triol are the kinetically preferred substrates in the case of HvUGT13248 and Bradi5g03300. We glucosylated several trichothecenes with OsUGT79 (HT-2 toxin, T-2 triol) and HvUGT13248 (T-2 toxin, neosolaniol, 4,15-diacetoxyscirpenol, fusarenon X) in the preparative scale. NMR analysis of the purified glucosides showed that exclusively β-D-glucosides were formed regio-selectively at position C-3-OH of the trichothecenes. These synthesized standards can be used to investigate the occurrence and toxicological properties of these modified mycotoxins.

Immunochemical Analysis of Paxilline and Ergot Alkaloid Mycotoxins in Grass Seeds and Plants.

PubMed

Bauer, Julia I; Gross, Madeleine; Cramer, Benedikt; Humpf, Hans-Ulrich; Hamscher, Gerd; Usleber, Ewald

2018-01-10

Limited availability of toxin standards for lolitrem B and ergovaline impedes routine control of grasses for endophyte toxins. This study aimed at assessing the applicability of an enzyme immunoassay (EIA) for the indole-diterpene mycotoxin paxilline, in combination with a generic EIA for ergot alkaloids, as alternative parameters for screening purposes. Analysis of grass seeds and model pastures of four different grass species showed that both EIAs yielded highly positive results for paxilline and ergot alkaloids in perennial ryegrass seeds. Furthermore, evidence for natural occurrence of paxilline in grass in Germany was obtained. High performance liquid chromatography-tandem mass spectrometry analysis qualitatively confirmed the paxilline EIA results but showed that paxilline analogues 1'-O-acetylpaxilline and 13-desoxypaxilline were the predominant compounds in seeds and grass. In the absence of easily accessible reference standards for specific analysis of some major endophyte toxins, analysis of paxilline and ergot alkaloids by EIA may be suitable substitute parameters. The major advantage of this approach is its ease of use and speed, providing an analytical tool which could enhance routine screening for endophyte toxins in pasture.

Volatile compounds of Aspergillus strains with different abilities to produce ochratoxin A.

PubMed

Jeleń, Henryk H; Grabarkiewicz-Szczesna, Jadwiga

2005-03-09

Volatile compounds emitted by Aspergillus strains having different abilities to produce ochratoxin A were investigated. Thirteen strains of Aspergillus ochraceus, three belonging to the A. ochraceus group, and eight other species of Aspergillus were examined for their abilities to produce volatile compounds and ochratoxin A on a wheat grain medium. The profiles of volatile compounds, analyzed using SPME, in all A. ochraceus strains, regardless of their toxeginicity, were similar and comprised mainly of 1-octen-3-ol, 3-octanone, 3-octanol, 3-methyl-1-butanol, 1-octene, and limonene. The prevailing compound was always 1-octen-3-ol. Mellein, which forms part of the ochratoxin A molecule, was found in both toxigenic and nontoxigenic strains. Volatile compounds produced by other Aspergillus strains were similar to those of A. ochraceus. Incubation temperatures (20, 24, and 27 degrees C) and water content in the medium (20, 30, and 40%) influenced both volatile compounds formation and ochratoxin A biosynthesis efficiency, although conditions providing the maximum amount of volatiles were different from those providing the maximum amount of ochratoxin A. The pattern of volatiles produced by toxigenic A. ochraceus strains does not facilitate their differentiation from nontoxigenic strains.

The history of ergot of rye (Claviceps purpurea) III: 1940-80.

PubMed

Lee, M R

2010-03-01

The period 1940-80 in the history of ergot was dominated by two investigators, Arthur Stoll and Albert Hofmann. There was great excitement when their group isolated from ergot preparations the powerful psychotropic agent lysergic acid diethylamide (LSD). It was thought that this substance would help to find the cause of schizophrenia and other psychotic disorders, but it would prove to be a great disappointment and Hofmann would say later, in private, that he regretted having spent so much time on the compound. By contrast, bromocriptine, derived from ergocriptine, would prove a pivotal substance in our knowledge of dopamine receptors in the central nervous system. It is widely used for the suppression of lactation, the treatment of prolactinomas and the management of Parkinson's disease.

Allelochemical regulation of reproduction and seed germination of two BrazilianBaccharis species by phytotoxic trichothecenes.

PubMed

Kuti, J O; Jarvis, B B; Mokhtari-Rejali, N; Bean, G A

1990-12-01

The potent phytotoxic trichothecene roridins and baccharinoids occur naturally in the Brazilian plants,Baccharis coridifolia andB. megapotamica. Biosynthesis of roridins inB. coridifolia appears to be linked to pollination, and the phytotoxins then accumulate in the seed. The roles of the phytotoxins in pollination, seed maturation, and germination of theBaccharis species were investigated. The high production of roridins occurred only in seeds resulting from intraspecific pollination, and the concentration of the toxins in the seeds generally increased with seed maturity. Removal of seed coats from trichothecene-producing BrazilianBaccharis species (B. coridifolia andB. megapotamica) and non-trichothecene-producing AmericanBaccharis species (B. halimifolia andB. glutinosa) resulted in improved seed germination ofB. halimifolia andB. glutinosa but complete inhibition of seed germination ofB. coridifolia andB. megapotamica. Addition of seed coat extracts of the BrazilianBaccharis species of dilute solutions (10(-6)μg/ml) of roridins or baccharinoids to the decoated seeds ofB. coridifolia andB. megapotamica resulted in germination, while seeds ofB. halimifolia andB. glutinosa were killed by the phytotoxins. Roridins interacted with gibberellic acid, a germination promoter, but not with abscisic acid, a germination inhibitor. The results from this study suggest that macrocyclic trichothecenes have a regulatory role(s) on reproduction and germination of BrazilianBaccharis species in their natural habitat.

Functional analysis of the gene controlling hydroxylation of festuclavine in the ergot alkaloid pathway of Neosartorya fumigata.

PubMed

Bilovol, Yulia; Panaccione, Daniel G

2016-11-01

Bioactive ergot alkaloids produced by several species of fungi are important molecules in agriculture and medicine. Much of the ergot alkaloid pathway has been elucidated, but a few steps, including the gene controlling hydroxylation of festuclavine to fumigaclavine B, remain unsolved. Festuclavine is a key intermediate in the fumigaclavine branch of the ergot alkaloid pathway of the opportunistic pathogen Neosartorya fumigata and also in the dihydrolysergic acid-based ergot alkaloid pathway of certain Claviceps species. Based on several lines of evidence, the N. fumigata gene easM is a logical candidate to encode the festuclavine-hydroxylating enzyme. To test this hypothesis we disrupted easM function by replacing part of its coding sequences with a hygromycin resistance gene and transforming N. fumigata with this construct. High-pressure liquid chromatography analysis demonstrated that easM deletion mutants were blocked in the ergot alkaloid pathway at festuclavine, and downstream products were eliminated. An additional alkaloid, proposed to be a prenylated form of festuclavine on the basis of mass spectral data, also accumulated to higher concentrations in the easM knockout. Complementation with the wild-type allele of easM gene restored the ability of the fungus to produce downstream compounds. These results indicate that easM encodes an enzyme required for fumigaclavine B synthesis likely by hydroxylating festuclavine. The festuclavine-accumulating strain of N. fumigata may facilitate future investigations of the biosynthesis of dihydrolysergic acid derivatives, which are derived from festuclavine and are the basis for several important drugs.

Biological detoxification of fungal toxins and its use in plant breeding, feed and food production.

PubMed

Karlovsky, P

1999-01-01

Enzymatic inactivation of fungal toxins is an attractive strategy for the decontamination of agricultural commodities and for the protection of crops from phytotoxic effects of fungal metabolites. This review summarizes research on the biological detoxification of fungal toxins by microorganisms and plants and its practical applications. Some mycotoxins are detoxified during ensiling and other fermentation processes (aflatoxins, alternariol, mycophenolic acid, patulin, PR toxin) while others are transformed into toxic products or survive fermentation unchanged. Plants can detoxify fomannoxin, fusaric acid, HC-toxin, ochratoxin A and oxalate but the degradation of deoxynivalenol has yet to be proven. Microflora of the digestive tract of vertebrates and invertebrates exhibit detoxification activities towards aflatoxins, ochratoxin A, oxalate and trichothecenes. Some toxin-producing fungi are able to degrade or transform their own products under suitable conditions. Pure cultures of bacteria and fungi which detoxify mycotoxins have been isolated from complex microbial populations by screening and enrichment culture techniques. Genes responsible for some of the detoxification activities have been cloned and expressed in heterologous hosts. The detoxification of aflatoxins, cercosporin, fumonisins, fusaric acid, ochratoxin A, oxalic acid, patulin, trichothecenes and zearalenone by pure cultures is reviewed. Finally, current application of these results in food and feed production and plant breeding is summarized and expected future developments are outlined. Copyright 1999 John Wiley & Sons, Ltd.

Screening and optimization of some inorganic salts for the production of ergot alkaloids from Penicillium species using surface culture fermentation process.

PubMed

Shahid, Memuna Ghafoor; Nadeem, Muhammad; Baig, Shahjehan; Cheema, Tanzeem Akbar; Atta, Saira; Ghafoor, Gul Zareen

2016-03-01

The present study deals with the production of ergot alkaloids from Penicillium commune and Penicillium citrinum, using surface culture fermentation process. Impact of various inorganic salts was tested on the production of ergot alkaloids during the optimization studies of fermentation medium such as impact of various concentration levels of succinic acid, ammonium chloride, MgSO4, FeSO4, ZnSO4, pH and the effect of various incubation time periods was also determined on the production of ergot alkaloids from Penicillium commune and Penicillium citrinum. Highest yield of ergot alkaloids was obtained when Penicillium commune and Penicillium citrinum that were grown on optimum levels of ingredients such as 2 g succinic acid, 1.5 and 2 g NH4Cl, 1.5 g MgSO4, 1 g FeSO4, 1 and 1.5 g ZnSO4 after 21 days of incubation time period using pH 5 at 25(°)C incubation temperature in the fermentation medium. Ergot alkaloids were determined using Spectrophotometry and Thin Layer Chromatography (TLC) techniques.

[Isolation of the ergot (Claviceps purpurea (Fr.) Tul., strain VKM-F-366D), producing the lactamic alkaloid ergocornam].

PubMed

Komarova, E L; Shain, S S; Sheĭchenko, V I

2002-01-01

A new ergot strain VKM-F-3662D producing lactamic alkaloid ergocornam with concomitant alkaloids valinamide and ergometrine was isolated during selective works with sclerotium MS-462, which was obtained from ergocryptine ergot strain VKM-F-2642D. The structure of these alkaloids was determined by 1H and 13C NMR.

Functional analysis of the gene controlling hydroxylation of festuclavine in the ergot alkaloid pathway of Neosartorya fumigata

PubMed Central

Bilovol, Yulia; Panaccione, Daniel G.

2016-01-01

Bioactive ergot alkaloids produced by several species of fungi are important molecules in agriculture and medicine. Much of the ergot alkaloid pathway has been elucidated, but a few steps, including the gene controlling hydroxylation of festuclavine to fumigaclavine B, remain unsolved. Festuclavine is a key intermediate in the fumigaclavine branch of the ergot alkaloid pathway of the opportunistic pathogen Neosartorya fumigata and also in the dihydrolysergic acid-based ergot alkaloid pathway of certain Claviceps species. Based on several lines of evidence, the N. fumigata gene easM is a logical candidate to encode the festuclavine-hydroxylating enzyme. To test this hypothesis we disrupted easM function by replacing part of its coding sequences with a hygromycin resistance gene and transforming N. fumigata with this construct. High pressure liquid chromatography analysis demonstrated that easM deletion mutants were blocked in the ergot alkaloid pathway at festuclavine, and downstream products were eliminated. An additional alkaloid, proposed to be a prenylated form of festuclavine on the basis of mass spectral data, also accumulated to higher concentrations in the easM knockout. Complementation with the wild-type allele of easM gene restored the ability of the fungus to produce downstream compounds. These results indicate that easM encodes an enzyme required for fumigaclavine B synthesis likely by hydroxylating festuclavine. The festuclavine-accumulating strain of N. fumigata may facilitate future investigations of the biosynthesis of dihydrolysergic acid derivatives, which are derived from festuclavine and are the basis for several important drugs. PMID:26972831

New insights into mycotoxin mixtures: The toxicity of low doses of Type B trichothecenes on intestinal epithelial cells is synergistic

DOE Office of Scientific and Technical Information (OSTI.GOV)

Alassane-Kpembi, Imourana; Université de Toulouse, ENVT, INP, UMR 1331 Toxalim, F-31076 Toulouse; Institut des Sciences Biomédicales Appliquées, Cotonou, Bénin

Deoxynivalenol (DON) is the most prevalent trichothecene mycotoxin in crops in Europe and North America. DON is often present with other type B trichothecenes such as 3-acetyldeoxynivalenol (3-ADON), 15-acetyldeoxynivalenol (15-ADON), nivalenol (NIV) and fusarenon-X (FX). Although the cytotoxicity of individual mycotoxins has been widely studied, data on the toxicity of mycotoxin mixtures are limited. The aim of this study was to assess interactions caused by co-exposure to Type B trichothecenes on intestinal epithelial cells. Proliferating Caco-2 cells were exposed to increasing doses of Type B trichothecenes, alone or in binary or ternary mixtures. The MTT test and neutral red uptake,more » respectively linked to mitochondrial and lysosomal functions, were used to measure intestinal epithelial cytotoxicity. The five tested mycotoxins had a dose-dependent effect on proliferating enterocytes and could be classified in increasing order of toxicity: 3-ADON < 15-ADON ≈ DON < NIV ≪ FX. Binary or ternary mixtures also showed a dose-dependent effect. At low concentrations (cytotoxic effect between 10 and 30–40%), mycotoxin combinations were synergistic; however DON–NIV–FX mixture showed antagonism. At higher concentrations (cytotoxic effect around 50%), the combinations had an additive or nearly additive effect. These results indicate that the simultaneous presence of low doses of mycotoxins in food commodities and diet may be more toxic than predicted from the mycotoxins alone. Considering the frequent co-occurrence of trichothecenes in the diet and the concentrations of toxins to which consumers are exposed, this synergy should be taken into account. - Highlights: • We assessed the individual and combined cytotoxicity of five trichothecenes. • The tested concentrations correspond to the French consumer exposure levels. • The type of interaction in combined cytotoxicity varied with the effect level. • Low doses of Type B trichothecenes induced

Microbial Inhibition of Fusarium Pathogens and Biological Modification of Trichothecenes in Cereal Grains

PubMed Central

Wachowska, Urszula; Packa, Danuta

2017-01-01

Fungi of the genus Fusarium infect cereal crops during the growing season and cause head blight and other diseases. Their toxic secondary metabolites (mycotoxins) contaminate grains. Several dozen toxic compounds produced by fungal pathogens have been identified to date. Type B trichothecenes—deoxynivalenol, its acetyl derivatives and nivalenol (produced mainly by F. graminearum and F. culmorum)—are most commonly detected in cereal grains. “T-2 toxin” (produced by, among others, F. sporotrichioides) belongs to type-A trichothecenes which are more toxic than other trichothecenes. Antagonistic bacteria and fungi can affect pathogens of the genus Fusarium via different modes of action: direct (mycoparasitism or hyperparasitism), mixed-path (antibiotic secretion, production of lytic enzymes) and indirect (induction of host defense responses). Microbial modification of trichothecenes involves acetylation, deacetylation, oxidation, de-epoxidation, and epimerization, and it lowers the pathogenic potential of fungi of the genus Fusarium. Other modifing mechanisms described in the paper involve the physical adsorption of mycotoxins in bacterial cells and the conjugation of mycotoxins to glucose and other compounds in plant and fungal cells. The development of several patents supports the commercialization and wider application of microorganisms biodegrading mycotoxins in grains and, consequently, in feed additives. PMID:29261142

Novel Zn(II)2Cys6 transcription factor in Fusarium equiseti trichothecene gene cluster is a pathway specific and global regulator

USDA-ARS?s Scientific Manuscript database

Trichothecenes are among the mycotoxins of greatest concern to food and feed safety and are produced by at least two lineages of Fusarium: the F. sambucinum (FSAMSC) and F. incarnatum-equiseti (FIESC) species complexes. Trichothecene biosynthesis begins with the formation of a cyclic sesquiterpene f...

Ergotism in Norway. Part 1: The symptoms and their interpretation from the late Iron Age to the seventeenth century.

PubMed

Alm, Torbjørn; Elvevåg, Brita

2013-03-01

Ergotism is a horrendous disease with grotesque symptoms caused by ingesting specific ergot alkaloids. Mass poisoning episodes are attributable to consumption of grain - usually rye - infected with the fungus Claviceps purpurea. By focusing on possible cases of ergotism, we re-examine Norwegian history from the sagas through to the end of the seventeenth century. Our review - not intended to be exhaustive, or ex post facto to assign medical or psychiatric labels - draws attention to the very real possibility that many remarkable medical cases may have been the result of the ingestion of highly poisonous and psychoactive food substances. Where possible we highlight explanations given at the time - often rooted in religion or demonology - to explain the disease.

Mitigation of ergot vasoconstriction by clover isoflavones in goats (Capra hircus)

USDA-ARS?s Scientific Manuscript database

Ergot alkaloids produced by a fungal endophyte (Epichloë coenophiala; formerly Neotyphodium coenphialum) that infects tall fescue (Lolium arundinaceum) can induce persistent constriction of the vasculature in ruminants, hindering their capability to thermo-regulate core body temperature. There is e...

Teratogenicity of Ochratoxin A and the Degradation Product, Ochratoxin α, in the Zebrafish (Danio rerio) Embryo Model of Vertebrate Development

PubMed Central

Haq, Mehreen; Gonzalez, Nelson; Mintz, Keenan; Jaja-Chimedza, Asha; De Jesus, Christopher Lawrence; Lydon, Christina; Welch, Aaron Z.; Berry, John P.

2016-01-01

Ochratoxins, and particularly ochratoxin A (OTA), are toxic fungal-derived contaminants of food and other agricultural products. Growing evidence supports the degradation of OTA by chemical, enzymatic and/or microbial means as a potential approach to remove this mycotoxin from food products. In particular, hydrolysis of OTA to ochratoxin α (OTα) and phenylalanine is the presumptive product of degradation in most cases. In the current study, we employed the zebrafish (Danio rerio) embryo, as a model of vertebrate development to evaluate, the teratogenicity of OTA and OTα. These studies show that OTA is potently active in the zebrafish embryo toxicity assay (ZETA), and that toxicity is both concentration- and time-dependent with discernible and quantifiable developmental toxicity observed at nanomolar concentrations. On the other hand, OTα had no significant effect on embryo development at all concentrations tested supporting a decreased toxicity of this degradation product. Taken together, these results suggest that ZETA is a useful, and highly sensitive, tool for evaluating OTA toxicity, as well as its degradation products, toward development of effective detoxification strategies. Specifically, the results obtained with ZETA, in the present study, further demonstrate the toxicity of OTA, and support its degradation via hydrolysis to OTα as an effective means of detoxification. PMID:26861395

Teratogenicity of Ochratoxin A and the Degradation Product, Ochratoxin α, in the Zebrafish (Danio rerio) Embryo Model of Vertebrate Development.

PubMed

Haq, Mehreen; Gonzalez, Nelson; Mintz, Keenan; Jaja-Chimedza, Asha; De Jesus, Christopher Lawrence; Lydon, Christina; Welch, Aaron; Berry, John P

2016-02-05

Ochratoxins, and particularly ochratoxin A (OTA), are toxic fungal-derived contaminants of food and other agricultural products. Growing evidence supports the degradation of OTA by chemical, enzymatic and/or microbial means as a potential approach to remove this mycotoxin from food products. In particular, hydrolysis of OTA to ochratoxin α (OTα) and phenylalanine is the presumptive product of degradation in most cases. In the current study, we employed the zebrafish (Danio rerio) embryo, as a model of vertebrate development to evaluate, the teratogenicity of OTA and OTα. These studies show that OTA is potently active in the zebrafish embryo toxicity assay (ZETA), and that toxicity is both concentration- and time-dependent with discernible and quantifiable developmental toxicity observed at nanomolar concentrations. On the other hand, OTα had no significant effect on embryo development at all concentrations tested supporting a decreased toxicity of this degradation product. Taken together, these results suggest that ZETA is a useful, and highly sensitive, tool for evaluating OTA toxicity, as well as its degradation products, toward development of effective detoxification strategies. Specifically, the results obtained with ZETA, in the present study, further demonstrate the toxicity of OTA, and support its degradation via hydrolysis to OTα as an effective means of detoxification.

Isolation and identification of trichothecenes from Fusarium compactum suspected in the aetiology of a major intoxication of sandhill cranes.

USGS Publications Warehouse

Cole, Richard J.; Dorner, Joe W.; Gilbert, John; Mortimer, David N.; Crews, Colin; Mitchell, J.C.; Windingstad, Ronald M.; Nelson, Paul E.; Cutler, Horace G.

1988-01-01

Isoneosolaniol (4,8-diacetoxy-12,13-epoxytrichothec-9-ene-3,15-diol) and other unidentified trichothecene mycotoxins were isolated from culture extracts of two highly toxigenic strains of Fusarium compactum cultured from waste peanuts involved in an acute intoxication of sandhill cranes (Grus canadensis). Neosolaniol and other unidentified trichothecenes were detected in waste peanuts collected from affected areas. The structure of isoneosolaniol was determined by 1H and 13C NMR analyses and by high-resolution mass spectometry. Isoneosolaniol was hightly toxic to 1-day-old chickens and to a HEp2 cell culture assay. It was concluded that the most logical cause of the sandhill crane intoxication was Fusarium spp. Contaminated peanuts and various trichothecene mycotoxins acting alone or in conjunction with other Fusarium mycotoxins.

Activities and effects of ergot alkaloids on livestock physiology and production

USDA-ARS?s Scientific Manuscript database

Ergot alkaloids can have a broad impact on many different physiological mechanisms that can alter the homeostasis of livestock exposed to these toxins through consumption of infested feedstuffs. This altered homeostasis causes an increased sensitivity in livestock to perturbations in the ambient env...

Purification, immunotoxic effects, and cellular uptake of trichothecene mycotoxins

DOE Office of Scientific and Technical Information (OSTI.GOV)

Witt, M.F.

1989-01-01

Studies were carried out to better understand how the trichothecenes alter immune function in animals and humans. Deoxynivalenol (DON) was purified for use in animal feeding studies. Dietary exposure to DON for 8 weeks altered the serum immunoglobulin profile in mice and decreased the splenic plaque-forming cell response to the antigen sheep red blood cells. The uptake of ({sup 3}H)T-2 toxin by a murine B-cell hybridoma was studied in order to learn more about the way in which trichothecenes interact with immune cells. A simple procedure was developed for the laboratory production and purification of gram quantities of crystalline DON.more » When Fusarium graminearum R6576 was grown on rice, concentrations of 600 to 700 ppm DON accumulated after 13 to 18 days of incubation. A DON derivative, 15-acetylDON, was also found at concentrations of 100 to 300 ppm after 7 to 10 days. DON was purified from crude culture extracts by water-saturated silica gel chromatography. Alpha-({sup 3}H)T-2 toxin of 99% chemical and radiochemical purity was prepared for use in uptake studies. Both the rate of uptake of ({sup 3}H)T-2 toxin by hybridomas and the time required for accumulation of ({sup 3}H)T-2 to reach equilibrium were proportional to the concentration of ({sup 3}H)T-2. ({sup 3}H)T-2 toxin accumulated by hybridomas was proportional to the concentration of ({sup 3}H)T-2 between 10{sup {minus}8} and 10{sup {minus}3} M. The rate of uptake of ({sup 3}H)jT-2 toxin by hybridomas was inhibited by the trichothecenes T-2 toxin, DON, verrucarin A, and roridin A, as well as the antibiotic anisomycin. The kinetics and concentration dependence of accumulation, along with the inhibition patterns, suggest that uptake of ({sup 3}H)T-2 toxin by hybridomas is mediated by binding of toxin to ribosomes.« less

Evaluation of Ochratoxin Recognition by Peptides Using Explicit Solvent Molecular Dynamics

PubMed Central

Thyparambil, Aby A.; Bazin, Ingrid; Guiseppi-Elie, Anthony

2017-01-01

Biosensing platforms based on peptide recognition provide a cost-effective and stable alternative to antibody-based capture and discrimination of ochratoxin-A (OTA) vs. ochratoxin-B (OTB) in monitoring bioassays. Attempts to engineer peptides with improved recognition efficacy require thorough structural and thermodynamic characterization of the binding-competent conformations. Classical molecular dynamics (MD) approaches alone do not provide a thorough assessment of a peptide’s recognition efficacy. In this study, in-solution binding properties of four different peptides, a hexamer (SNLHPK), an octamer (CSIVEDGK), NFO4 (VYMNRKYYKCCK), and a 13-mer (GPAGIDGPAGIRC), which were previously generated for OTA-specific recognition, were evaluated using an advanced MD simulation approach involving accelerated configurational search and predictive modeling. Peptide configurations relevant to ochratoxin binding were initially generated using biased exchange metadynamics and the dynamic properties associated with the in-solution peptide–ochratoxin binding were derived from Markov State Models. Among the various peptides, NFO4 shows superior in-solution OTA sensing and also shows superior selectivity for OTA vs. OTB due to the lower penalty associated with solvating its bound complex. Advanced MD approaches provide structural and energetic insights critical to the hapten-specific recognition to aid the engineering of peptides with better sensing efficacies. PMID:28505090

Ochratoxin A on green coffee: influence of harvest and drying processing procedures.

PubMed

Paulino De Moraes, Maria Heloisa; Luchese, Rosa Helena

2003-09-10

Ochratoxin A is a metabolite produced by Aspergillus and Penicillium species that is nephrotoxic and possibly carcinogenic to humans. The aim of this study was to evaluate ochratoxin A contamination in green coffee obtained by different harvesting and drying operations and from fruits of different ripening stages in order to identify hazards. The research was directed to coffees from the highland area of Rio de Janeiro state (Brazil), which is traded in the domestic market. Twenty-two out of 54 samples contained ochratoxin A at levels ranging from 0.3 to 160 microg/kg. Ochatoxin A contamination levels between different ripe stage fruits were not significant (P > 0.05). "Varrição" coffee, consisting of fruits that fell from the tree spontaneously and stayed longer on the ground before being harvested, was the most contaminated. Eleven out of 14 samples of varrição coffee were contaminated. Three out of 10 samples from the northwestern region of the state were positive for ochratoxin at levels ranging from 10.1 to 592 microg/kg. The contaminated samples had in common the fact that they were harvested directly from the soil.

Effect of acute exposure to ergot alkaloids on short-chain fatty acid absorption and barrier function of isolated bovine ruminal epithelium

USDA-ARS?s Scientific Manuscript database

Ergot alkaloids present in endophyte-infected tall fescue are the causative agents for fescue toxicosis in cattle. Ergot alkaloids have been shown to cause a reduction in blood flow to the rumen epithelium as well as a decrease in short-chain fatty acid (SCFA) absorption from the washed rumen of ste...

Trichothecene 3-O-acetyltransferase protects both the producing organism and transformed yeast from related mycotoxins. Cloning and characterization of Tri101.

PubMed

Kimura, M; Kaneko, I; Komiyama, M; Takatsuki, A; Koshino, H; Yoneyama, K; Yamaguchi, I

1998-01-16

Trichothecene mycotoxins such as deoxynivalenol, 4,15-diacetoxyscirpenol, and T-2 toxin, are potent protein synthesis inhibitors for eukaryotic organisms. The 3-O-acetyl derivatives of these toxins were shown to reduce their in vitro activity significantly as assessed by assays using a rabbit reticulocyte translation system. The results suggested that the introduction of an O-acetyl group at the C-3 position in the biosynthetic pathway works as a resistance mechanism for Fusarium species that produce t-type trichothecenes (trichothecenes synthesized via the precursor trichotriol). A gene responsible for the 3-O-acetylation reaction, Tri101, has been successfully cloned from a Fusarium graminearum cDNA library that was designed to be expressed in Schizosaccharomyces pombe. Fission yeast transformants were selected for their ability to grow in the presence of T-2 toxin, and this strategy allowed isolation of 25 resistant clones, all of which contained a cDNA for Tri101. This is the first drug-inactivating O-acetyltransferase gene derived from antibiotic-producing organisms. The open reading frame of Tri101 codes for a polypeptide of 451 amino acid residues, which shows no similarity to any other proteins reported so far. TRI101 from recombinant Escherichia coli catalyzes O-acetylation of the trichothecene ring specifically at the C-3 position in an acetyl-CoA-dependent manner. By using the Tri101 cDNA as a probe, two least overlapping cosmid clones that cover a region of 70 kilobase pairs have been isolated from the genome of F. graminearum. Other trichothecene biosynthetic genes, Tri4, Tri5, and Tri6, were not clustered in the region covered by these cosmid clones. These new cosmid clones are considered to be located in other parts of the large biosynthetic gene cluster and might be useful for the study of trichothecene biosynthesis.

The ergot alkaloid gene cluster in Claviceps purpurea: extension of the cluster sequence and intra species evolution.

PubMed

Haarmann, Thomas; Machado, Caroline; Lübbe, Yvonne; Correia, Telmo; Schardl, Christopher L; Panaccione, Daniel G; Tudzynski, Paul

2005-06-01

The genomic region of Claviceps purpurea strain P1 containing the ergot alkaloid gene cluster [Tudzynski, P., Hölter, K., Correia, T., Arntz, C., Grammel, N., Keller, U., 1999. Evidence for an ergot alkaloid gene cluster in Claviceps purpurea. Mol. Gen. Genet. 261, 133-141] was explored by chromosome walking, and additional genes probably involved in the ergot alkaloid biosynthesis have been identified. The putative cluster sequence (extending over 68.5kb) contains 4 different nonribosomal peptide synthetase (NRPS) genes and several putative oxidases. Northern analysis showed that most of the genes were co-regulated (repressed by high phosphate), and identified probable flanking genes by lack of co-regulation. Comparison of the cluster sequences of strain P1, an ergotamine producer, with that of strain ECC93, an ergocristine producer, showed high conservation of most of the cluster genes, but significant variation in the NRPS modules, strongly suggesting that evolution of these chemical races of C. purpurea is determined by evolution of NRPS module specificity.

Ergot alkaloids: From witchcraft till in silico analysis. Multi-receptor analysis of ergotamine metabolites.

PubMed

Dellafiora, Luca; Dall'Asta, Chiara; Cozzini, Pietro

2015-01-01

The term Ergot is referred to the sclerotium of ascomycetes - a protective kernel produced during resting stage of some fungi - which replaces seeds of susceptible cereals and plants intended for human and animal diet. It contains various composition of tryptophan-derived toxins defined ergot alkaloids. Since sclerotia can be harvested and milled together with cereals, they represent a source of food and feed contamination after breakage and spreading of mycotoxins into the various milling fractions. The effects of ergot alkaloids, including those adverse for human health, have been known since the Middle Ages. Nevertheless, as recently stated by the European Food Safety Authority, further information is needed on metabolism and target receptors-binding of common alkaloids in food. Unfortunately, the experimental investigation is challenging due to the high costs in terms of time and money. This study was thus aimed at assessing whether the in silico modeling can be an effective tool to investigate the interaction between multiple serotonin receptors and a wide set of ergotamine metabolites, including experimentally detected molecules and predicted derivatives. Validated models provided precious insights about the effects exerted by metabolic modifications on the receptor-ligand interaction. Such structural information may be useful to support the design of further experimental analysis.

Production of ochratoxin A by Aspergillus ochraceus NRRL-3174 before and after exposures to 60Co irradiation.

PubMed Central

Applegate, K L; Chipley, J R

1976-01-01

Spores from the toxigenic organism Aspergillus ochraceus NRRL-3174 were exposed to specific levels of gamma irradiation and then allowed to germinate on selected media. Increases in ochratoxin A production by irradiated, compared to non-irradiated, spores were observed after inoculation of spores onto a cracked red wheat or into a synthetic liquid medium. Variations in daily ochratoxin production were also observed for control and irradiated spore-derived cultures developing on both media, with maximum toxin production varying from 7 to 11 days of incubation. The most notable increases in ochratoxin A production occurred from cultures developing from spores having been irradiated with 10, 25, or 50 krad. Exposures to 400 or 600 krad resulted in complete inhibition of spore germination and, consequently, no ochratoxin production. Of the two substrates used, wheat and synthetic, the quantities of ochratoxin A produced were significantly lower in the synthetic media than on the natural substrate. Higher and more rapid toxin production occurred from spores having been irradiated with 10, 25, 50, and 100 krad than occurred from the non-irradiated control spores when grown on synthetic media. Cultures derived from spores having been exposed to 10, 25, 50, and 100 krad produced significantly higher levels of ochratoxin A after 8 days of incubation on natural substrate than did the controls. Analysis of variance revealed that substrate, length of incubation, as well as irradiation levels all affected the time required to produce maximum levels of ochratoxin A. PMID:938031

The Statement that Some Ochratoxins are “..classified as human carcinogens” is Not Accurate

USDA-ARS?s Scientific Manuscript database

To the Editor, We are writing in response to a statement made in the recent “Editor’s Highlights” of the paper on ochratoxin and the microbiome (Toxicological Sciences, 141(1), 2014, 1). Ochratoxins, like the other agriculturally-important mycotoxins, are natural contaminants and must be managed o...

Mechanisms of Action of Trichothecenes in the Cardiovascular System

DTIC Science & Technology

1986-10-01

L.: Stachybotryotoxicosis as a veterinary problem. In Mycotoxins in Human and Animal Health . (Rodricks, J.V., Hesseltine, C.W. and Mehlman, M.A., eds...trichothecenes. In Myocotoxins in Human anti Animal Health . (Rodricks, J.V., Hosseltine, C.W. and Mehlman, M.A., 2ds.), Pathotox Publishers, Inc., Park...2i. Pier, A.C.: Mycotcxins and Animal Health . Advances in Veterinary Science and Comparative Medicine 25:185-243, 1981. / 22. Sato, N. and Ueno, Y

Fate of trichothecene mycotoxins during the processing: milling and baking.

PubMed

Lancova, K; Hajslova, J; Kostelanska, M; Kohoutkova, J; Nedelnik, J; Moravcova, H; Vanova, M

2008-05-01

Toxic secondary metabolites produced by fungi representing Fusarium genus are common contaminants in cereals worldwide. To estimate the dietary intake of these trichothecene mycotoxins, information on their fate during cereal processing is needed. Up-to-date techniques such as high-performance liquid chromatography (HPLC) coupled with tandem mass spectrometry (LC-MS/MS) was used for the analysis of seven trichothecenes (deoxynivalenol, nivalenol, HT-2 toxin, T-2 toxin, 15- and 3-acetyldeoxynivalenol, and fusarenon-X) in bread production chain (wheat grains, intermediate products collected during milling and baking process, breads). Regardless of whether the grains were naturally infected or artificially inoculated by Fusarium spp. in the field, the fractions obtained from the grain-cleaning procedure contained the highest mycotoxin levels. During milling the highest concentrations of deoxynivalenol were found in the bran, the lowest in the reduction flours. Baking at 210 degrees C for 14 min had no significant effect on deoxynivalenol levels. The rheological properties of dough measured by fermentograph, maturograph, oven rise recorder, and laboratory baking test were carried out, and based on the obtained results the influence of mycotoxin content on rheological behaviour was investigated.

Ergot alkaloids reduce rumen epithelial blood flow and volatile fatty acid absorption

USDA-ARS?s Scientific Manuscript database

Ergot alkaloids have been shown to induce vasoconstriction of both peripheral and ruminal vessels. Constriction of ruminal vessels could lead to a reduction in epithelial blood flow thereby reducing nutrient absorption. The objectives of this experiment were to determine if steers receiving endophyt...

Fumonisin and Ochratoxin Production in Industrial Aspergillus niger Strains

PubMed Central

Frisvad, Jens C.; Larsen, Thomas O.; Thrane, Ulf; Meijer, Martin; Varga, Janos; Samson, Robert A.; Nielsen, Kristian F.

2011-01-01

Aspergillus niger is perhaps the most important fungus used in biotechnology, and is also one of the most commonly encountered fungi contaminating foods and feedstuffs, and occurring in soil and indoor environments. Many of its industrial applications have been given GRAS status (generally regarded as safe). However, A. niger has the potential to produce two groups of potentially carcinogenic mycotoxins: fumonisins and ochratoxins. In this study all available industrial and many non-industrial strains of A. niger (180 strains) as well as 228 strains from 17 related black Aspergillus species were examined for mycotoxin production. None of the related 17 species of black Aspergilli produced fumonisins. Fumonisins (B2, B4, and B6) were detected in 81% of A. niger, and ochratoxin A in 17%, while 10% of the strains produced both mycotoxins. Among the industrial strains the same ratios were 83%, 33% and 26% respectively. Some of the most frequently used strains in industry NRRL 337, 3112 and 3122 produced both toxins and several strains used for citric acid production were among the best producers of fumonisins in pure agar culture. Most strains used for other biotechnological processes also produced fumonisins. Strains optimized through random mutagenesis usually maintained their mycotoxin production capability. Toxigenic strains were also able to produce the toxins on media suggested for citric acid production with most of the toxins found in the biomass, thereby questioning the use of the remaining biomass as animal feed. In conclusion it is recommended to use strains of A. niger with inactive or inactivated gene clusters for fumonisins and ochratoxins, or to choose isolates for biotechnological uses in related non-toxigenic species such as A. tubingensis, A. brasiliensis, A vadensis or A. acidus, which neither produce fumonisins nor ochratoxins. PMID:21853139

In silico analysis sheds light on the structural basis underlying the ribotoxicity of trichothecenes-A tool for supporting the hazard identification process.

PubMed

Dellafiora, Luca; Galaverna, Gianni; Dall'Asta, Chiara

2017-03-15

Deoxynivalenol is a food borne mycotoxin belonging to the trichothecenes family that may cause severe injuries in human and animals. The inhibition of protein synthesis via the interaction with the ribosome has been identified as a crucial mechanism underlying toxic action. However, it is not still fully understood how and to what extent compounds belonging to trichothecenes family affect human and animal health. In turn, this scenario causes delay in managing the related health risk. Aimed at supporting the hazard identification process, the in silico analysis may be a straightforward tool to investigate the structure-activity relationship of trichothecenes, finding out molecules of possible concern to carry forth in the risk assessment process. In this framework, this work investigated through a molecular modeling approach the structural basis underlying the interaction with the ribosome under a structure-activity relationship perspective. To identify further forms possibly involved in the total trichothecenes-dependent ribotoxic load, the model was challenged with a set of 16 trichothecene modified forms found in plants, fungi and animals, including also compounds never tested before for the capability to bind and inhibit the ribosome. Among them, only the regiospecific glycosylation in the position 3 of the sesquiterpenoid scaffold (i.e. T-2 toxin-3-glucuronide, α and β isomers of T-2 toxin-3-glucoside and deoxynivalenol-3-glucuronide) was found impairing the interaction with the ribosome, while the other compounds tested (i.e. neosolaniol, nivalenol, fusarenon-X, diacetoxyscirpenol, NT-1 toxin, HT-2 toxin, 19- and 20-hydroxy-T-2 toxin, T-2 toxin triol and tetraol, and 15-deacetyl-T-2 toxin), were found potentially able to inhibit the ribosome. Accordingly, they should be included with high priority in further risk assessment studies in order to better characterize the trichothecenes-related hazard. Copyright © 2017 Elsevier B.V. All rights reserved.

Evolutionary history of ergot with a new infrageneric classification (Hypocreales: Clavicipitaceae: Claviceps).

PubMed

Píchová, Kamila; Pažoutová, Sylvie; Kostovčík, Martin; Chudíčková, Milada; Stodůlková, Eva; Novák, Petr; Flieger, Miroslav; van der Linde, Elna; Kolařík, Miroslav

2018-06-01

The ergot, genus Claviceps, comprises approximately 60 species of specialised ovarial grass parasites famous for the production of food toxins and pharmaceutics. Although the ergot has been known for centuries, its evolution have not been resolved yet. Our approach combining multilocus phylogeny, molecular dating and the study of ecological, morphological and metabolic features shows that Claviceps originated in South America in the Palaeocene on a common ancestor of BEP (subfamilies Bambusoideae, Ehrhartoideae, Pooideae) and PACMAD (subfamilies Panicoideae, Aristidoideae, Chloridoideae, Micrairoideae, Arundinoideae, Danthonioideae) grasses. Four clades described here as sections diverged during the Paleocene and Eocene. Since Claviceps are parasitic fungi with a close relationship with their host plants, their evolution is influenced by interactions with the new hosts, either by the spread to a new continent or the radiation of the host plants. Three of the sections possess very narrow host ranges and biogeographical distributions and have relatively low toxicity. On the contrary, the section Claviceps, comprising the rye ergot, C. purpurea, is unique in all aspects. Fungi in this section of North American origin have spread all over the world and infect grasses in all subfamilies as well as sedges, and it is the only section synthesising toxic ergopeptines and secalonic acids. The evolutionary success of the Claviceps section members can be explained by high toxin presence, serving as feeding deterrents and playing a role in their protective mutualism with host plants. Closely related taxa Neoclaviceps monostipa and Cepsiclava phalaridis were combined into the genus Aciculosporium. Copyright © 2018 Elsevier Inc. All rights reserved.

Extraction of ochratoxin A in red wine with dopamine-coated magnetic multi-walled carbon nanotubes.

PubMed

Wan, Hong; Zhang, Bo; Bai, Xiao-Lin; Zhao, Yan; Xiao, Meng-Wei; Liao, Xun

2017-10-01

A new, rapid, green, and cost-effective magnetic solid-phase extraction of ochratoxin A from red wine samples was developed using polydopamine-coated magnetic multi-walled carbon nanotubes as the absorbent. The polydopamine-coated magnetic multi-walled carbon nanotubes were fabricated with magnetic multi-walled carbon nanotubes and dopamine by an in situ oxidative self-polymerization approach. Transmission electron microscopy, dynamic light scattering, X-ray photoelectron spectroscopy and vibrating sample magnetometry were used to characterize the absorbents. Ochratoxin A was quantified with high-performance liquid chromatography coupled with fluorescence detection, with excitation and emission wavelengths of 338 and 455 nm, respectively. The conditions affecting the magnetic solid-phase extraction procedure, such as pH, extraction solution, extraction time, absorbent amount, desorption solution and desorption time were investigated to obtain the optimal extraction conditions. Under the optimized conditions, the extraction recovery was 91.8-104.5% for ochratoxin A. A linear calibration curve was obtained in the range of 0.1-2.0 ng/mL. The limit of detection was 0.07 ng/mL, and the limit of quantitation was 0.21 ng/mL. The recoveries of ochratoxin A for spiked red wine sample ranged from 95.65 to 100.65% with relative standard deviation less than 8%. The polydopamine-coated magnetic multi-walled carbon nanotubes showed a high affinity toward ochratoxin A, allowing selective extraction and quantification of ochratoxin A from complex sample matrixes. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

New insights into the organization and regulation of trichothecene biosynthetic genes in Trichoderma

USDA-ARS?s Scientific Manuscript database

Collectively, species of the genus Trichoderma can produce numerous structurally diverse secondary metabolites (SM). This ability is conferred by the presence of SM biosynthetic gene clusters in their genomes. Species of Trichoderma in the Brevicompactum clade are able to produce trichothecenes, a f...

The identification of QTL controlling ergot sclerotia size in hexaploid wheat implicates a role for the Rht dwarfing alleles.

PubMed

Gordon, Anna; Basler, Ryan; Bansept-Basler, Pauline; Fanstone, Vicky; Harinarayan, Lakshmi; Grant, Paul K; Birchmore, Richard; Bayles, Rosemary A; Boyd, Lesley A; O'Sullivan, Donal M

2015-12-01

Four QTL conferring resistance to ergot were identified in the UK winter wheat varieties 'Robigus' and 'Solstice'. Two QTL co-located with semi-dwarfing alleles at the Rht loci Rht - 1B and Rht - 1D implicating a role of these DELLA proteins in infection success of Claviceps purpurea. The fungal pathogen Claviceps purpurea infects ovaries of a broad range of temperate grasses and cereals, including hexaploid wheat, causing a disease commonly known as ergot. Sclerotia produced in place of seed carry a cocktail of harmful alkaloid compounds that result in a range of symptoms in humans and animals, causing ergotism. Following a field assessment of C. purpurea infection in winter wheat, two varieties 'Robigus' and 'Solstice' were selected which consistently produced the largest differential effect on ergot sclerotia weights. They were crossed to produce a doubled haploid mapping population, and a marker map, consisting of 714 genetic loci and a total length of 2895 cM was produced. Four ergot reducing QTL were identified using both sclerotia weight and size as phenotypic parameters; QCp.niab.2A and QCp.niab.4B being detected in the wheat variety 'Robigus', and QCp.niab.6A and QCp.niab.4D in the variety 'Solstice'. The ergot resistance QTL QCp.niab.4B and QCp.niab.4D peaks mapped to the same markers as the known reduced height (Rht) loci on chromosomes 4B and 4D, Rht-B1 and Rht-D1, respectively. In both cases, the reduction in sclerotia weight and size was associated with the semi-dwarfing alleles, Rht-B1b from 'Robigus' and Rht-D1b from 'Solstice'. Two-dimensional, two-QTL scans identified significant additive interactions between QTL QCp.niab.4B and QCp.niab.4D, and between QCp.niab.2A and QCp.niab.4B when looking at sclerotia size, but not between QCp.niab.2A and QCp.niab.4D. The two plant height QTL, QPh.niab.4B and QPh.niab.4D, which mapped to the same locations as QCp.niab.4B and QCp.niab.4D, also displayed significant genetic interactions.

Antibody binding of circulating ergot alkaloids in cattle grazing tall fescue.

PubMed

Hill, N S; Thompson, F N; Dawe, D L; Stuedemann, J A

1994-03-01

Direct evidence linking alkaloids found in endophyte-infected tall fescue forage with the livestock disorder known as fescue toxicosis is lacking. Physiologic effects of fescue toxicosis include reduced serum prolactin concentration in cattle. A monoclonal antibody specific to the lysergic moiety of ergot alkaloids was developed in mice after creating an immunogen by linking lysergol to human serum albumin. The antibody was specific to the lysergic moiety and, therefore, it cross-reacted with ergot alkaloids, lysergic acid, and lysergol. The antibody did not cross-react with alkaloid derivatives that had bromated or hydrogenated lysergic ring moieties. Fescue toxicosis conditions were elicited in yearling Angus steers by permitting them to graze endophyte-infected tall fescue containing > 650 micrograms/kg of ergovaline for 60 days. Passive immunization of steers by infusion of the monoclonal antibody increased serum prolactin concentration by 7 ng/ml, beginning immediately after infusion. Control steers did not respond to treatment with bovine serum albumin. Active immunization of yearling Angus heifers with immunogens containing lysergol or ergonovine linked to human serum albumin resulted in an antibody response.

Application of Liquid Chromatography/Ion Trap Mass Spectrometry Technique to Determine Ergot Alkaloids in Grain Products

PubMed Central

Szymczyk, Krystyna; Jędrzejczak, Renata; Roszko, Marek

2015-01-01

Summary A liquid chromatography/ion trap mass spectrometry-based method to determine six ergot alkaloids and their isomers is presented. The samples were cleaned on neutral alumina-based solid-phase extraction cartridges. The following method parameters were obtained (depending on the analyte and spiking level): method recovery from 63.0 to 104.6%, relative standard deviation below 18%, linear range from 1 to 325 µg/kg, linear correlation coefficient not less than 0.98. The developed analytical procedure was applied to determine the levels of ergot alkaloids in 65 samples of selected rye-based food products (flour – 34 samples, bran – 12 samples, rye – 18 samples, flakes – 1 sample). Measurable levels of alkaloids were found in majority of the analysed samples, particularly in rye flour. Additionally, alkaloids were determined in ergot sclerotia isolated from rye grains. Total content was nearly 0.01% (97.9 mg/kg). However, the alkaloid profile was dominated by ergocristine at 45.6% (44.7 mg/kg), an alkaloid not commonly found in the tested food products. Ergocorninine at 0.2% (0.2 mg/kg) was the least abundant alkaloid. PMID:27904328

Physiologic effects of ergot alkaloids: What happens when excretion does not equal absorption?

USDA-ARS?s Scientific Manuscript database

Increased persistence of tall fescue (Lolium arundinaceum) infested with an endophytic fungus Epichloë coenophiala (formerly Neotyphodium coenophialum) in forage-based agriculture has led to increased effort in understanding the negative effects caused by consumption of ergot alkaloids by animals co...

Physiologic effects of ergot alkaloids: What happens when excretion does not equal consumption?

USDA-ARS?s Scientific Manuscript database

Increased persistence of tall fescue (Lolium arundinaceum) infested with an endophytic fungus Epichloë coenophiala (formerly Neotyphodium coenophialum) in forage-based agriculture has led to increased effort in understanding the negative effects caused by consumption of ergot alkaloids by animals co...

Comments on "Ochratoxin A: In utero Exposure in Mice Induces Adducts in Testicular DNA. Toxins 2010, 2, 1428-1444"-Mis-Citation of Rat Literature to Justify a Hypothetical Role for Ochratoxin A in Testicular Cancer.

PubMed

Mantle, Peter G

2010-10-01

A manuscript in the journal recently cited experimental rat data from two manuscripts to support plausibility of a thesis that ochratoxin A might be a cause of human testicular cancer. I believe that there is no experimental evidence that ochratoxin A produces testicular cancer in rats or mice.

Spatio-temporal dynamics of Fusarium head blight and Trichothecene toxin types in Canada

USDA-ARS?s Scientific Manuscript database

In many parts of the world Fusarium graminearum is the primary causal agent of Fusarium head blight (FHB), a disease of cereal crops that adversely affects crop yield, food safety, and animal health. We previously demonstrated population structure associated with differences in trichothecene toxin t...

Biosynthesis of the Pharmaceutically Important Fungal Ergot Alkaloid Dihydrolysergic Acid Requires a Specialized Allele of cloA

PubMed Central

Arnold, Stephanie L.

2017-01-01

ABSTRACT Ergot alkaloids are specialized fungal metabolites that are important as the bases of several pharmaceuticals. Many ergot alkaloids are derivatives of lysergic acid (LA) and have vasoconstrictive activity, whereas several dihydrolysergic acid (DHLA) derivatives are vasorelaxant. The pathway to LA is established, with the P450 monooxygenase CloA playing a key role in oxidizing its substrate agroclavine to LA. We analyzed the activities of products of cloA alleles from different fungi relative to DHLA biosynthesis by expressing them in a mutant of the fungus Neosartorya fumigata that accumulates festuclavine, the precursor to DHLA. Transformants expressing CloA from Epichloë typhina × Epichloë festucae, which oxidizes agroclavine to LA, failed to oxidize festuclavine to DHLA. In substrate feeding experiments, these same transformants oxidized exogenously supplied agroclavine to LA, indicating that a functional CloA was produced. A genomic clone of cloA from Claviceps africana, a sorghum ergot fungus that produces a DHLA derivative, was cloned and expressed in the festuclavine-accumulating mutant of N. fumigata, but several introns in this genomic clone were not processed properly. Expression of a synthetic intron-free version of C. africana cloA resulted in the accumulation of DHLA as assessed by fluorescence high-pressure liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LC-MS). In substrate feeding experiments, the C. africana CloA also accepted agroclavine as the substrate, oxidizing it to LA. The data indicate that a specialized allele of cloA is required for DHLA biosynthesis and that the pharmaceutically important compound DHLA can be produced in engineered N. fumigata. IMPORTANCE Ergot alkaloids are fungal metabolites that have impacted humankind historically as poisons and more recently as pharmaceuticals used to treat dementia, migraines, and other disorders. Much is known about the biosynthesis of ergot alkaloids that are

Biosynthesis of the Pharmaceutically Important Fungal Ergot Alkaloid Dihydrolysergic Acid Requires a Specialized Allele of cloA.

PubMed

Arnold, Stephanie L; Panaccione, Daniel G

2017-07-15

Ergot alkaloids are specialized fungal metabolites that are important as the bases of several pharmaceuticals. Many ergot alkaloids are derivatives of lysergic acid (LA) and have vasoconstrictive activity, whereas several dihydrolysergic acid (DHLA) derivatives are vasorelaxant. The pathway to LA is established, with the P450 monooxygenase CloA playing a key role in oxidizing its substrate agroclavine to LA. We analyzed the activities of products of cloA alleles from different fungi relative to DHLA biosynthesis by expressing them in a mutant of the fungus Neosartorya fumigata that accumulates festuclavine, the precursor to DHLA. Transformants expressing CloA from Epichloë typhina × Epichloë festucae , which oxidizes agroclavine to LA, failed to oxidize festuclavine to DHLA. In substrate feeding experiments, these same transformants oxidized exogenously supplied agroclavine to LA, indicating that a functional CloA was produced. A genomic clone of cloA from Claviceps africana , a sorghum ergot fungus that produces a DHLA derivative, was cloned and expressed in the festuclavine-accumulating mutant of N. fumigata , but several introns in this genomic clone were not processed properly. Expression of a synthetic intron-free version of C. africana cloA resulted in the accumulation of DHLA as assessed by fluorescence high-pressure liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LC-MS). In substrate feeding experiments, the C. africana CloA also accepted agroclavine as the substrate, oxidizing it to LA. The data indicate that a specialized allele of cloA is required for DHLA biosynthesis and that the pharmaceutically important compound DHLA can be produced in engineered N. fumigata IMPORTANCE Ergot alkaloids are fungal metabolites that have impacted humankind historically as poisons and more recently as pharmaceuticals used to treat dementia, migraines, and other disorders. Much is known about the biosynthesis of ergot alkaloids that are

Trichothecene resistance in wheat: Development of molecular markers for PDR-type ABC transporter genes.

PubMed

Mitterbauer, R; Heinrich, M; Rauscher, R; Lemmens, M; Bürstmayr, H; Adam, G

2003-03-01

Infection withFusarium graminearum andF. culmorum not only causes severe yield and quality losses, the most relevant concern is the contamination of cereal foods and feeds with trichothecenes (e.g. deoxynivalenol, DON). The ability to synthesize trichothecenes has been shown to be a virulence factor ofF. graminearum on wheat and, on the other hand, toxin resistance is most likely an important component of field resistance. Our hypothesis is that pleiotropic drug resistance mediated by PDR-type ABC transporter proteins (acting as membrane located drug efflux pumps) is a relevant mechanism of DON resistance not only in yeast but also in wheat. Goal of this project is the development of molecular markers for this gene family for use in marker-assisted plant breeding programs. The technical difficulties caused by the large size of the PDR-family are discussed.

Comparison of Anorectic Potencies of the Trichothecenes T-2 Toxin, HT-2 Toxin and Satratoxin G to the Ipecac Alkaloid Emetine.

PubMed

Wu, Wenda; Zhou, Hui-Ren; Pan, Xiao; Pestka, James J

Trichothecene mycotoxins, potent translational inhibitors that are associated with human food poisonings and damp-building illnesses, are of considerable concern to animal and human health. Food refusal is a hallmark of exposure of experimental animals to deoxynivalenol (DON) and other Type B trichothecenes but less is known about the anorectic effects of foodborne Type A trichothecenes (e.g., T-2 toxin, HT-2 toxin), airborne Type D trichothecenes (e.g. satratoxin G [SG]) or functionally analogous metabolites that impair protein synthesis. Here, we utilized a well-described mouse model of food intake to compare the anorectic potencies of T-2 toxin, HT-2 toxin, and SG to that of emetine, a medicinal alkaloid derived from ipecac that inhibits translation. Intraperitoneal (IP) administration with T-2 toxin, HT-2 toxin, emetine and SG evoked anorectic responses that occurred within 0.5 h that lasted up to 96, 96, 3 and 96 h, respectively, with lowest observed adverse effect levels (LOAELs) being 0.1, 0.1, 2.5 and 0.25 mg/kg BW, respectively. When delivered via natural routes of exposure, T-2 toxin, HT-2 toxin, emetine (oral) and SG (intranasal) induced anorectic responses that lasted up to 48, 48, 3 and 6 h, respectively with LOAELs being 0.1, 0.1, 0.25, and 0.5 mg/kg BW, respectively. All four compounds were generally much more potent than DON which was previously observed to have LOAELs of 1 and 2.5 mg/kg BW after IP and oral dosing, respectively. Taken together, these anorectic potency data will be valuable in discerning the relative risks from trichothecenes and other translational inhibitors of natural origin.

Incorporation of Chlorine-36 into Ochratoxin A

PubMed Central

Wei, Ru-Dong; Strong, F. M.; Smalley, E. B.

1971-01-01

Incubation of Aspergillus ochraceus NRRL 3174 in a medium containing Na36Cl incorporated 36Cl into ochratoxin A. The highest incorporation (0.75 and 0.70%, respectively) was obtained when the radioactive chloride was added to the medium by the 2nd or 3rd day of incubation. Images PMID:5119199

Phytotoxicity Evaluation of Type B Trichothecenes Using a Chlamydomonas reinhardtii Model System

PubMed Central

Suzuki, Tadahiro; Iwahashi, Yumiko

2014-01-01

Type B trichothecenes, which consist of deoxynivalenol (DON) and nivalenol (NIV) as the major end products, are produced by phytotoxic fungi, such as the Fusarium species, and pollute arable fields across the world. The DON toxicity has been investigated using various types of cell systems or animal bioassays. The evaluation of NIV toxicity, however, has been relatively restricted because of its lower level compared with DON. In this study, the Chlamydomonas reinhardtii testing system, which has been reported to have adequate NIV sensitivity, was reinvestigated under different mycotoxin concentrations and light conditions. The best concentration of DON and NIV, and their derivatives, for test conditions was found to be 25 ppm (2.5 × 10−2 mg/mL). In all light test conditions, DON, NIV, and fusarenon-X (FusX) indicated significant growth inhibition regardless of whether a light source existed, or under differential wavelength conditions. FusX growth was also influenced by changes in photon flux density. These results suggest that C. reinhardtii is an appropriate evaluation system for type B trichothecenes. PMID:24476708

Variation in the expression of ergot alkaloids between individual tillers of perennial ryegrass

NASA Astrophysics Data System (ADS)

Mace, Wade; Lunn, Kristy; Lloyd-West, Catherine

2014-11-01

Epichloë fungal endophytes of cool season grasses are well known to produce a range of alkaloids of benefit to the host. Some of these compounds are advantageous to agriculture due to qualities that promote pasture persistence (e.g. the loline class of alkaloids confer insect protection) while others are detrimental to the wellbeing of grazing livestock. The ergot alkaloids (e.g. ergovaline), produced in ryegrass and tall fescue associations, causes poor animal health in farming regions in many countries around the world and further study is required to improve our knowledge on this class of compounds. Here we present the application of a quantitative LC-MS/MS (liquid chromatography coupled to mass spectrometry) method measuring eight ergot alkaloids (chanoclavine, agroclavine, elymoclavine, lysergol, lysergic acid, ergine, lysergyl alanine, ergovaline) produced by endophyte infected grasses, to monitor levels in individual tillers from multiple plants of a single cultivar of perennial ryegrass (Lolium perenne cv. ‘Grasslands Samson’) infected with a common toxic endophyte strain (Epichloë festucae var. lolii). Monitoring the expression in individual tillers allows an estimation of the variability within a plant (between tillers) as well as between plants. The study showed that there is significant variation in the concentration of the ergot alkaloids between tillers of a single plant, at or exceeding the level of variation observed between individual plants of a population. This result emphasizes the fundamental importance of robust experimental design and sampling procedures when alkaloid expression assessment is required and these need to be rigorously tailored to the hypothesis being tested.

Convergent and divergent evolution of the trichothecene mycotoxin biosynthetic gene cluster in the Fusarium

USDA-ARS?s Scientific Manuscript database

The Fusarium trichothecenes nivalenol (NIV) and deoxynivalenol (DON) are among the mycotoxins of greatest concern to agricultural production and food/feed safety worldwide. Previous analyses indicate that during early evolution of the Fusarium incarnatum-F. equiseti species complex (FIESC), the tri...

Physiological responses to known intake of ergot alkaloids by steers at environmental temperatures within or greater than their thermoneutral zone

NASA Astrophysics Data System (ADS)

Eisemann, Joan; Huntington, Gerald; Williamson, Megan; Hanna, Michelle; Poore, Matthew

2014-11-01

Two studies separated effects of dietary ergot alkaloids from effects of feed intake or ambient temperature on respiration rate (RR), heart rate (HR), surface temperature (ST), rectal temperature (RT), blood pressure (BP), serum hormone, and plasma metabolite concentrations in beef steers. The balanced, single reversal design for each experiment used 8 beef steers fed tall fescue seed (2.5 g/kg body weight, (BW)) with (E+) or without (E-) ergot alkaloids as part of a 60:40 switchgrass hay: supplement diet. Periods were 35 d with 21 d of preliminary phase and 14 d of feeding fescue seed once daily. Measures of dependent variables were collected on d 20, 25, 29 and 35 of each period at 0730 (before feeding), 1230 and 1530. In Expt 1 steers weighed 286 kg, gained 0.61 kg BW/d, E+ supplied 2.72 mg ergot alkaloids including 1.60 mg ergovaline per steer daily, and mean minimum and maximum daily ambient temperatures were 23.6 and 32.3°C. In Expt 2 steers weighed 348 kg, gained 1.03 kg BW/d, E+ supplied 3.06 mg ergot alkaloids including 2.00 mg ergovaline daily, and mean minimum and maximum daily ambient temperatures were 11.9 and 17.4°C. Dry matter intake was not affected by fescue seed treatment (P < 0.20) in either experiment. In both experiments, E+ reduced HR (P < 0.01) and increased insulin (P = 0.07). Systolic BP minus diastolic BP decreased (P< 0.05) for E+ in both experiments, due to increased diastolic BP in Expt 1 (P < 0.03) and decreased systolic BP in Expt 2 (P < 0.07). In Expt 1, above the thermoneutral zone, E+ increased (P< 0.05) RR, RT and left side ST in comparison to E-, but in Expt 2, within the thermoneutral zone, E+ and E- did not differ (P < 0.18). Ergot alkaloids from fescue seed affect the cardiovascular system of steers separately from effects of feed intake or environmental temperature. Ergot alkaloids interact with ambient temperatures above the steers’ thermoneutral zone to exacerbate the symptoms of hyperthermic stress.

A Water-Damaged Home and Health of Occupants: A Case Study

PubMed Central

Thrasher, Jack Dwayne; Gray, Michael R.; Kilburn, Kaye H.; Dennis, Donald P.; Yu, Archie

2012-01-01

A family of five and pet dog who rented a water-damaged home and developed multiple health problems. The home was analyzed for species of mold and bacteria. The diagnostics included MRI for chronic sinusitis with ENT and sinus surgery, and neurological testing for neurocognitive deficits. Bulk samples from the home, tissue from the sinuses, urine, nasal secretions, placenta, umbilical cord, and breast milk were tested for the presence of trichothecenes, aflatoxins, and Ochratoxin A. The family had the following diagnosed conditions: chronic sinusitis, neurological deficits, coughing with wheeze, nose bleeds, and fatigue among other symptoms. An infant was born with a total body flare, developed multiple Cafe-au-Lait pigmented skin spots and diagnoses with NF1 at age 2. The mycotoxins were detected in bulk samples, urine and nasal secretions, breast milk, placenta, and umbilical cord. Pseudomonas aueroginosa, Acinetobacter, Penicillium, and Aspergillus fumigatus were cultured from nasal secretions (father and daughter). RT-PCR revealed A. fumigatus DNA in sinus tissues of the daughter. The dog had 72 skin lesions (sebaceous glands and lipomas) from which trichothecenes and ochratoxin A. were detected. The health of the family is discussed in relation to the most recent published literature regarding microbial contamination and toxic by-products present in water-damaged buildings. PMID:22220187

A water-damaged home and health of occupants: a case study.

PubMed

Thrasher, Jack Dwayne; Gray, Michael R; Kilburn, Kaye H; Dennis, Donald P; Yu, Archie

2012-01-01

A family of five and pet dog who rented a water-damaged home and developed multiple health problems. The home was analyzed for species of mold and bacteria. The diagnostics included MRI for chronic sinusitis with ENT and sinus surgery, and neurological testing for neurocognitive deficits. Bulk samples from the home, tissue from the sinuses, urine, nasal secretions, placenta, umbilical cord, and breast milk were tested for the presence of trichothecenes, aflatoxins, and Ochratoxin A. The family had the following diagnosed conditions: chronic sinusitis, neurological deficits, coughing with wheeze, nose bleeds, and fatigue among other symptoms. An infant was born with a total body flare, developed multiple Cafe-au-Lait pigmented skin spots and diagnoses with NF1 at age 2. The mycotoxins were detected in bulk samples, urine and nasal secretions, breast milk, placenta, and umbilical cord. Pseudomonas aueroginosa, Acinetobacter, Penicillium, and Aspergillus fumigatus were cultured from nasal secretions (father and daughter). RT-PCR revealed A. fumigatus DNA in sinus tissues of the daughter. The dog had 72 skin lesions (sebaceous glands and lipomas) from which trichothecenes and ochratoxin A. were detected. The health of the family is discussed in relation to the most recent published literature regarding microbial contamination and toxic by-products present in water-damaged buildings.

Assessment of the electronic structure and properties of trichothecene toxins using density functional theory

USDA-ARS?s Scientific Manuscript database

A comprehensive quantum chemical study was carried out on 34 type A and type B trichothecenes, including selected derivatives and biosynthetic precursors of deoxynivalenol, nivalenol, and T-2 toxin. Quantum parameters, Natural Bond Orbital (NBO) analysis, and molecular properties were calculated on ...

Quantitative molecular diagnostic assays of grain washes for Claviceps purpurea are correlated with visual determinations of ergot contamination.

PubMed

Comte, Alexia; Gräfenhan, Tom; Links, Matthew G; Hemmingsen, Sean M; Dumonceaux, Tim J

2017-01-01

We examined the epiphytic microbiome of cereal grain using the universal barcode chaperonin-60 (cpn60). Microbial community profiling of seed washes containing DNA extracts prepared from field-grown cereal grain detected sequences from a fungus identified only to Class Sordariomycetes. To identify the fungal sequence and to improve the reference database, we determined cpn60 sequences from field-collected and reference strains of the ergot fungus, Claviceps purpurea. These data allowed us to identify this fungal sequence as deriving from C. purpurea, and suggested that C. purpurea DNA is readily detectable on agricultural commodities, including those for which ergot was not identified as a grading factor. To get a sense of the prevalence and level of C. purpurea DNA in cereal grains, we developed a quantitative PCR assay based on the fungal internal transcribed spacer (ITS) and applied it to 137 samples from the 2014 crop year. The amount of Claviceps DNA quantified correlated strongly with the proportion of ergot sclerotia identified in each grain lot, although there was evidence that non-target organisms were responsible for some false positives with the ITS-based assay. We therefore developed a cpn60-targeted loop-mediated isothermal amplification assay and applied it to the same grain wash samples. The time to positive displayed a significant, inverse correlation to ergot levels determined by visual ratings. These results indicate that both laboratory-based and field-adaptable molecular diagnostic assays can be used to detect and quantify pathogen load in bulk commodities using cereal grain washes.

Quantitative molecular diagnostic assays of grain washes for Claviceps purpurea are correlated with visual determinations of ergot contamination

PubMed Central

Comte, Alexia; Gräfenhan, Tom; Links, Matthew G.; Hemmingsen, Sean M.

2017-01-01

We examined the epiphytic microbiome of cereal grain using the universal barcode chaperonin-60 (cpn60). Microbial community profiling of seed washes containing DNA extracts prepared from field-grown cereal grain detected sequences from a fungus identified only to Class Sordariomycetes. To identify the fungal sequence and to improve the reference database, we determined cpn60 sequences from field-collected and reference strains of the ergot fungus, Claviceps purpurea. These data allowed us to identify this fungal sequence as deriving from C. purpurea, and suggested that C. purpurea DNA is readily detectable on agricultural commodities, including those for which ergot was not identified as a grading factor. To get a sense of the prevalence and level of C. purpurea DNA in cereal grains, we developed a quantitative PCR assay based on the fungal internal transcribed spacer (ITS) and applied it to 137 samples from the 2014 crop year. The amount of Claviceps DNA quantified correlated strongly with the proportion of ergot sclerotia identified in each grain lot, although there was evidence that non-target organisms were responsible for some false positives with the ITS-based assay. We therefore developed a cpn60-targeted loop-mediated isothermal amplification assay and applied it to the same grain wash samples. The time to positive displayed a significant, inverse correlation to ergot levels determined by visual ratings. These results indicate that both laboratory-based and field-adaptable molecular diagnostic assays can be used to detect and quantify pathogen load in bulk commodities using cereal grain washes. PMID:28257512

[Levels of Ochratoxin A and total Aflatoxins in Panamanian exportation coffee by an ELISA Method].

PubMed

Franco, Heriberto; Vega, Aracelly; Reyes, Stephany; De Léon, Javier; Bonilla, Alexis

2014-03-01

A study about processing conditions of exportation coffee in 15 benefits located in Chiriqui, western region of Panama, was conducted. In addition, 21 samples of processed coffee (green beans), from the benefits, were analyzed. The samples were microbiologically tested in order to quantify total aflatoxins (B1, B2, G1 and G2) and Ochratoxin A (OTA), using the immunoaffinity ELISA method. A detection limit of 0.017 ng/mL, was determined for Ochratoxin A, which is equivalent to a concentration of 0.829 µg/kg, and a detection limit of 0.027 ng/mL, for total aflatoxins, which is equivalent to a concentration of 1.350 µg/kg. It was found that four (19%) out of the 21 samples were positive to the presence of Ochratoxin A and three (14%) to the presence of total aflatoxins. Samples showed levels of Ochratoxin A in the range 4.90 - 37.73 µg/kg; only three of them exceeded the maximum limit allowed by the European Union, for the concentration of Ochratoxin, which is of 5.0 µg/kg. Total aflatoxins were found in the range 1.51 - 1.93 µg/kg, below 10 µg/kg which is the maximum limit allowed for coffee by the European Union. The results indicate that the processing of coffee produced in Panama successfully meets international standards for postharvest handling, which leads to a low incidence ofmycotoxins and very low levels ofmycotoxin-producing fungi.

Vasoconstrictive responses by the carotid and auricular arteries in goats to ergot alkaloid exposure

USDA-ARS?s Scientific Manuscript database

A fungal endophyte (Neotyphodium coenophialum) infects most plants of ‘Kentucky 31’ tall fescue (Lolium arundinaceum) and produces ergot alkaloids that cause persistent constriction of the vascular system in grazing livestock. Consequently, animals undergoing this toxicosis cannot regulate core body...

[Issenheim's altarpiece: does the disease depicted on the altarpiece correspond to ergotism?].

PubMed

Faure, Michel

2010-01-01

The Issenheim altar by Mathias Grünenwald (Unterlinden museum in Colmar) shows various scenes from the life of St Antony, abbot. The Tentation panel has been considered as an illustration of the sacer ignis, or St Antony's disease (ergotism). In fact, it may more likely indicate an early representation of Syphilis, or the French disease, as discussed here.

Electrospray[+] tandem quadrupole mass spectrometry in the elucidation of ergot alkaloids chromatographed by HPLC: screening of grass or forage samples for novel toxic compounds.

PubMed

Lehner, Andreas F; Craig, Morrie; Fannin, Neil; Bush, Lowell; Tobin, Tom

2005-11-01

Ergot alkaloids are mycotoxins generated by grass and grain pathogens such as Claviceps, for example. Ergot alkaloid-poisoning syndromes, such as tall fescue toxicosis from endophyte-infected tall fescue grass, are important veterinary problems for cattle, horses, sheep, pigs and chickens, with consequent impact on food, meat and dairy industries. Damage to livestock is of the order of a billion dollars a year in the United States alone. HPLC with UV and fluorescence detection are the predominant means of ergot alkaloid determination, with focus on quantitation of the marker compound ergovaline, although ELISA methods are undergoing investigation. These techniques are excellent for rapid detection, but of poor specificity in defining new or poorly characterized ergot alkaloids and related compounds. This paper demonstrates the facility of using electrospray(+) mass spectrometry with multiple reaction monitoring (MRM) detection during chromatographic examination of ergot alkaloid standards of lysergic acid, lysergol, ergonovine, ergovaline, ergotamine, ergocornine, ergocryptine and ergocrystine by HPLC. Ergoline-8 position epimers could be separated on the gradient HPLC system for ergocornine, ergocrystine and ergonovine and appeared as shoulders for ergotamine and ergovaline; epimers generally showed different patterns of relative intensity for specific MRM transitions. There was reasonable correspondence between retention of standards on the 2-mm ESI(+)MS phenyl-hexyl-based reverse phase column and those on the 4-mm C18-based column. Since up to 10% of clinical cases involving toxin exposure display unidentified chromatographic peaks, 11 samples of feed components associated with such cases were studied with developed MRM methods to attempt elucidation of crucial components if possible. Ergotamine appeared in all, ergovaline appeared in five and ergocornine appeared in six; ergonovine, ergocryptine, ergocrystine and lysergol also appeared in several. In addition

Lysergic acid amide as chemical marker for the total ergot alkaloids in rye flour - Determination by high-performance thin-layer chromatography-fluorescence detection.

PubMed

Oellig, Claudia

2017-07-21

Ergot alkaloids are generally determined by high-performance liquid chromatography (HPLC) coupled to fluorescence detection (FLD) or mass selective detection, analyzing the individual compounds. However, fast and easy screening methods for the determination of the total ergot alkaloid content are more suitable, since for monitoring only the sum of the alkaloids is relevant. The herein presented screening uses lysergic acid amide (LSA) as chemical marker, formed from ergopeptine alkaloids, and ergometrine for the determination of the total ergot alkaloids in rye with high-performance thin-layer chromatography-fluorescence detection (HPTLC-FLD). An ammonium acetate buffered extraction step was followed by liquid-liquid partition for clean-up before the ergopeptine alkaloids were selectively transformed to LSA and analyzed by HPTLC-FLD on silica gel with isopropyl acetate/methanol/water/25% ammonium hydroxide solution (80:10:3.8:1.1, v/v/v/v) as the mobile phase. The enhanced native fluorescence of LSA and unaffected ergometrine was used for quantitation without any interfering matrix. Limits of detection and quantitation were 8 and 26μg LSA/kg rye, which enables the determination of the total ergot alkaloids far below the applied quality criterion limit for rye. Close to 100% recoveries for different rye flours at relevant spiking levels were obtained. Thus, reliable results were guaranteed, and the fast and efficient screening for the total ergot alkaloids in rye offers a rapid alternative to the HPLC analysis of the individual compounds. Copyright © 2017 Elsevier B.V. All rights reserved.

[Determination of ochratoxin A by ELISA I. Study on the preparation of ochratoxin a antigen].

PubMed

Chen, Xuelan; Xu, Yang; Wu, Chenggang

2002-02-01

Ochratoxin A (OTA) antigen was prepared by activated ester method. Factors influencing OTA antigen were discussed and the optimum conditions was found by L9(3(4)) orthogonal design. The results showed that when the mole ratio was OTA:BSA = 20:1, OTA:NHS(N-hydroxy-succinamide):DCC(dicyclohexylcarbodiimide) = 1:2:4, the activate time was 120 min and the conjugation time was 90 min, the utilization of OTA could reach 48.2% and the best conjugation ratio of OTA and BSA was 9.64.

Wavelength-dependent degradation of ochratoxin and citrinin by light in vitro and in vivo and its implications on Penicillium.

PubMed

Schmidt-Heydt, Markus; Cramer, Benedikt; Graf, Irina; Lerch, Sandra; Humpf, Hans-Ulrich; Geisen, Rolf

2012-12-14

It has previously been shown that the biosynthesis of the mycotoxins ochratoxin A and B and of citrinin by Penicillium is regulated by light. However, not only the biosynthesis of these mycotoxins, but also the molecules themselves are strongly affected by light of certain wavelengths. The white light and blue light of 470 and 455 nm are especially able to degrade ochratoxin A, ochratoxin B and citrinin after exposure for a certain time. After the same treatment of the secondary metabolites with red (627 nm), yellow (590 nm) or green (530 nm) light or in the dark, almost no degradation occurred during that time indicating the blue light as the responsible part of the spectrum. The two derivatives of ochratoxin (A and B) are degraded to certain definitive degradation products which were characterized by HPLC-FLD-FTMS. The degradation products of ochratoxin A and B did no longer contain phenylalanine however were still chlorinated in the case of ochratoxin A. Citrinin is completely degraded by blue light. A fluorescent band was no longer visible after detection by TLC suggesting a higher sensitivity and apparently greater absorbance of energy by citrinin. The fact that especially blue light degrades the three secondary metabolites is apparently attributed to the absorption spectra of the metabolites which all have an optimum in the short wave length range. The absorption range of citrinin is, in particular, broader and includes the wave length of blue light. In wheat, which was contaminated with an ochratoxin A producing culture of Penicillium verrucosum and treated with blue light after a pre-incubation by the fungus, the concentration of the preformed ochratoxin A reduced by roughly 50% compared to the control and differed by > 90% compared to the sample incubated further in the dark. This indicates that the light degrading effect is also exerted in vivo, e.g., on food surfaces. The biological consequences of the light instability of the toxins are discussed.

Ergot species of the Claviceps purpurea group from South Africa.

PubMed

van der Linde, Elna J; Pešicová, Kamila; Pažoutová, Sylvie; Stodůlková, Eva; Flieger, Miroslav; Kolařík, Miroslav

2016-08-01

Results of a survey and study of the Claviceps purpurea group of species in South Africa are being presented and five new species are described. Morphological descriptions are based on the anamorphs and four nuclear genetic loci. Claviceps fimbristylidis sp. nov. on Fimbristylis complanata was discovered wide-spread across five provinces of the country associated with water and represents the fourth Claviceps species recorded from the Cyperaceae. Claviceps monticola sp. nov. is described from Brachypodium flexum growing in mountain forests in Mpumalanga Province, as well as the northern Drakensberg southwards into the Eastern Cape Province. Claviceps pazoutovae sp. nov. is recorded from Stipa dregeana var. dregeana and Ehrharta erecta var. erecta, also associated with these mountain ranges. Claviceps macroura sp. nov. is recorded from Cenchrus macrourus from the Eastern Cape and Claviceps capensis sp. nov. from Ehrharta villosa var. villosa is recorded from the Western Cape Province. Claviceps cyperi, only recorded from South Africa is included in the study. Ergot alkaloid profiles of all species are provided and showed similarity to C. purpurea. Only C. cyperi and in lesser degree C. capensis, C. macroura, and C. pazoutovae produced ergot alkaloids in clinically significant amounts. Several reported species infect invasive grass species, native to South Africa, and thus represent potentially invasive species. Copyright © 2016 British Mycological Society. Published by Elsevier Ltd. All rights reserved.

Comments on “Ochratoxin A: In utero Exposure in Mice Induces Adducts in Testicular DNA. Toxins 2010, 2, 1428–1444”—Mis-Citation of Rat Literature to Justify a Hypothetical Role for Ochratoxin A in Testicular Cancer

PubMed Central

Mantle, Peter G.

2010-01-01

A manuscript in the journal recently cited experimental rat data from two manuscripts to support plausibility of a thesis that ochratoxin A might be a cause of human testicular cancer. I believe that there is no experimental evidence that ochratoxin A produces testicular cancer in rats or mice. PMID:22069555

From the Lab Bench: Season Changes in Ergot Alkaloid Concentrations of Toxic Tall Fescue

USDA-ARS?s Scientific Manuscript database

A column was written to discuss seasonal trends in ergot alkaloids produced by the fungal endophyte that infects most plants of tall fescue. Tall fescue is the predominant grass utilized for forage in the U.S. transition zone between the temperature northeast and subtropical southeast; however, erg...

Validation and transferability study of a method based on near-infrared hyperspectral imaging for the detection and quantification of ergot bodies in cereals.

PubMed

Vermeulen, Ph; Fernández Pierna, J A; van Egmond, H P; Zegers, J; Dardenne, P; Baeten, V

2013-09-01

In recent years, near-infrared (NIR) hyperspectral imaging has proved its suitability for quality and safety control in the cereal sector by allowing spectroscopic images to be collected at single-kernel level, which is of great interest to cereal control laboratories. Contaminants in cereals include, inter alia, impurities such as straw, grains from other crops, and insects, as well as undesirable substances such as ergot (sclerotium of Claviceps purpurea). For the cereal sector, the presence of ergot creates a high toxicity risk for animals and humans because of its alkaloid content. A study was undertaken, in which a complete procedure for detecting ergot bodies in cereals was developed, based on their NIR spectral characteristics. These were used to build relevant decision rules based on chemometric tools and on the morphological information obtained from the NIR images. The study sought to transfer this procedure from a pilot online NIR hyperspectral imaging system at laboratory level to a NIR hyperspectral imaging system at industrial level and to validate the latter. All the analyses performed showed that the results obtained using both NIR hyperspectral imaging cameras were quite stable and repeatable. In addition, a correlation higher than 0.94 was obtained between the predicted values obtained by NIR hyperspectral imaging and those supplied by the stereo-microscopic method which is the reference method. The validation of the transferred protocol on blind samples showed that the method could identify and quantify ergot contamination, demonstrating the transferability of the method. These results were obtained on samples with an ergot concentration of 0.02% which is less than the EC limit for cereals (intervention grains) destined for humans fixed at 0.05%.

[Immunoaffinity columns and determination of ochratoxin A in cereals by HPLC. Part I.: Evaluation of extraction using methanol/water].

PubMed

Czerwiecki, Ludwik; Czyzyk, Kamila; Kwiecień, Agnieszka; Wilczyńska, Grazyna

2004-01-01

The method for ochratoxin A determination in cereals (wheat, rye) was described. Application of immunoaffinity columns (IAC) for clean-up of extracts was investigated. The ochratoxin A content was determined by high-performance liquid chromatography using C18 column and fluorimetric detection at 330 nm (excitation) and 460 nm (emission). The mean recovery of ochratoxin A was 65-78%. The limits of detection (LOD) and quantification (LOQ) were 0.015 and 0.025 microg/kg, respectively. The positive results were confirmed by reaction with BF3 complex in methanol.

Pathological Outcomes in Kidney and Brain in Male Fischer Rats Given Dietary Ochratoxin A, Commencing at One Year of Age

PubMed Central

Mantle, Peter G.; Nolan, Christopher C.

2010-01-01

Malignant renal carcinoma, manifest in morbid ageing rats, is the striking component of an otherwise silent response after about nine months of exposure to ochratoxin A in the first year of life (daily intake ~100-250 µg/kg body weight). Reasons for the long latency are unclear, as is whether there would be a similar carcinogenic response if toxin exposure started at one year of age. Therefore, 24 male Fischer rats were given 100 µg ochratoxin A as a daily dietary contaminant for 35 weeks from age 50 weeks. Plasma ochratoxin A concentration reached a maximum value of ~8 µg/mL within one month of starting the toxin regimen. No renal carcinomas occurred. Four renal adenomas, two of which were only microscopic, were found among the six rats surviving for 110 weeks. The findings raise new questions about a difference between young adults and mature adults in sensitivity of male rats to the ochratoxin A-induced DNA damage necessary for renal carcinogenesis. A pilot histological study of perfuse-fixed brains of the toxin-treated rats showed no gross abnormalities, correlating with the consistent absence of behavioral or neurological disorders from chronic ochratoxin A exposure regimens in the range 100-250 µg/kg/day during the second half of life. Reasoned questioning concerning ochratoxin A as a neurotoxic mycotoxin is made. PMID:22069628

Derivatives of Ergot-alkaloids: Molecular structure, physical properties, and structure-activity relationships

NASA Astrophysics Data System (ADS)

Ivanova, Bojidarka B.; Spiteller, Michael

2012-09-01

A comprehensive screening of fifteen functionalized Ergot-alkaloids, containing bulk aliphatic cyclic substituents at D-ring of the ergoline molecular skeleton was performed, studying their structure-active relationships and model interactions with α2A-adreno-, serotonin (5HT2A) and dopamine D3 (D3A) receptors. The accounted high affinity to the receptors binding loops and unusual bonding situations, joined with the molecular flexibility of the substituents and the presence of proton accepting/donating functional groups in the studied alkaloids, may contribute to further understanding the mechanisms of biological activity in vivo and in predicting their therapeutic potential in central nervous system (CNS), including those related the Schizophrenia. Since the presented correlation between the molecular structure and properties, was based on the comprehensively theoretical computational and experimental physical study on the successfully isolated derivatives, through using routine synthetic pathways in a relatively high yields, marked these derivatives as 'treasure' for further experimental and theoretical studied in areas such as: (a) pharmacological and clinical testing; (b) molecular-drugs design of novel psychoactive substances; (c) development of the analytical protocols for determination of Ergot-alkaloids through a functionalization of the ergoline-skeleton, and more.

Evidence that enforced sunlight exposure can cause hyperthermia in cattle ingesting low levels of ergot of rye (Claviceps purpurea), when air temperature and humidity conditions are only moderate.

PubMed

Bourke, C A

2003-09-01

To determine the rectal temperature response of cattle, following the oral administration of ergot of rye (Claviceps purpurea), under pen conditions of enforced sunlight compared with those of enforced shade. Hereford cross steers were divided into two groups of 18. One group was dosed once, on a Monday morning, with finely ground rye grass ergots at a rate of 180 mg/kg body weight and held in the sun for 7 h each day until Friday afternoon. The other group was not dosed but was similarly held in the sun during the same period. Their rectal temperatures were measured early morning and mid afternoon, from Monday to Friday inclusive. The process was repeated for each group, but this time they were held in the shade. The four treatment options were run concurrently by conducting the experiments over 6 weeks and using 3 animals in each treatment group, each week. The thermic response over all weeks, of the ergot treated, sunlight exposed cattle, was deemed greater than for the other groups, based on the following four parameters. The increase in rectal temperature between early morning and mid afternoon, the size of the mid afternoon rectal temperature rise, the difference between the maximum mid afternoon rectal temperature recorded by an animal in the sun compared with that recorded by the same animal in the shade, and finally the number of animals in a treatment group that recorded rectal temperatures > 40.00 degrees C. The difference in the daily increase in body temperature between the ergot treated, sun exposed cattle and the ergot treated, shaded cattle, was greater than that observed between the sun exposed and shade restricted control cattle. Nine of 18 ergot treated and sun exposed cattle developed hyperthermia; no cattle in the other three groups did. Some sunlight exposed cattle, dosed with a low amount of ergot of rye, can experience a body temperature elevation above the normal range, even under mild ambient temperature and humidity conditions. Sunlight

Production and characterization of monoclonal antibodies against ochratoxin B.

PubMed

Heussner, Alexandra H; Moeller, Ines; Day, Billy W; Dietrich, Daniel R; O'Brien, Evelyn

2007-05-01

Monoclonal antibodies against ochratoxin B (OTB) were generated by immunizing Balb/c mice with OTB conjugated to keyhole limpet hemocyanin (KLH) via carbodiimide reactions with CHMC and EDAC. A stable hybridoma cell line 2F1.E10 was produced by fusion of murine splenocytes and myeloma cells. The obtained antibodies were characterized using an indirect competitive ELISA. The detection limit was calculated (27+/-2 nM OTB) and 50% binding inhibition was reached at 500 nM free OTB. A low cross-reactivity to ochratoxin A (OTA) of 3.3% and no cross-reactivities to either coumarin or DL-phenylalanine were observed, suggesting a highly specific OTB antibody. The antibody type was identified as IgG class 1 with the light chain being of the kappa configuration. These antibodies can be used in an indirect competitive ELISA to detect OTB in the nanomolar to micromolar concentration range and may be useful for the analysis of contaminated food items.

Natural Phenolic Inhibitors of Trichothecene Biosynthesis by the Wheat Fungal Pathogen Fusarium culmorum: A Computational Insight into the Structure-Activity Relationship

PubMed Central

Pani, Giovanna; Dessì, Alessandro; Dallocchio, Roberto; Scherm, Barbara; Azara, Emanuela; Delogu, Giovanna

2016-01-01

A model of the trichodiene synthase (TRI5) of the wheat fungal pathogen and type-B trichothecene producer Fusarium culmorum was developed based on homology modelling with the crystallized protein of F. sporotrichioides. Eight phenolic molecules, namely ferulic acid 1, apocynin 2, propyl gallate 3, eugenol 4, Me-dehydrozingerone 5, eugenol dimer 6, magnolol 7, and ellagic acid 8, were selected for their ability to inhibit trichothecene production and/or fungal vegetative growth in F. culmorum. The chemical structures of phenols were constructed and partially optimised based on Molecular Mechanics (MM) studies and energy minimisation by Density Functional Theory (DFT). Docking analysis of the phenolic molecules was run on the 3D model of F. culmorum TRI5. Experimental biological activity, molecular descriptors and interacting-structures obtained from computational analysis were compared. Besides the catalytic domain, three privileged sites in the interaction with the inhibitory molecules were identified on the protein surface. The TRI5-ligand interactions highlighted in this study represent a powerful tool to the identification of new Fusarium-targeted molecules with potential as trichothecene inhibitors. PMID:27294666

Cytotoxicity of trichothecenes and fusarochromanone produced by Fusarium equiseti strains isolated from Norwegian cereals.

PubMed

Morrison, Ellen; Rundberget, Thomas; Kosiak, Barbara; Aastveit, Are H; Bernhoft, Aksel

2002-01-01

The cytotoxicity and secondary metabolites of 28 Norwegian strains of Fusarium equiseti have been characterized. Trichothecenes and fusarochromanone (FUCH) in rice culture extracts of the strains were analysed by gas chromatography-mass spectrometry (GC-MS) and high performance liquid chromatography (HPLC). The following metabolites were found in all isolates: FUCH, nivalenol (NIV), scirpentriol (SCIRP), 4-acetylnivalenol (4-ac-NIV, also called fusarenon-X), 15-acetyl-nivalenol (15-ac-NIV), and diacetoxyscirpenol (DAS). 4,15-diacetyl-nivalenol (diacetyl-NIV) was found in 5 isolates. Porcine kidney epithelial cells (PK15. American Type Culture Collection) were exposed to rice culture extracts to study cytotoxicity. Descriptive statistics and factor analysis of the identified secondary metabolites show that their main metabolites were FUCH, NIV, SCIRP, DAS and 15-ac-NIV, consecutively. The individual trichothecenes were highly intercorrelated, whereas the production of acetylated NIV and DAS was slightly less. Stepwise multiple regression analysis of cytotoxicity and metabolite profiles of rice culture extracts ascribed the toxicity mainly to a combination of FUCH and 15-ac-NIV, though SCIRP or DAS are agents in the combined toxicity as well.

New Insight into the Ochratoxin A Biosynthetic Pathway through Deletion of a Nonribosomal Peptide Synthetase Gene in Aspergillus carbonarius

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gallo, A.; Bruno, K. S.; Solfrizzo, M.

2012-09-14

Ochratoxin A (OTA), a mycotoxin produced by Aspergillus and Penicillium species, is composed of a dihydroisocoumarin ring linked to phenylalanine and its biosynthetic pathway has not yet been completely elucidated. Most of the knowledge regarding the genetic and enzymatic aspects of OTA biosynthesis has been obtained in Penicillium species. In Aspergillus species only pks genes involved in the initial steps of the pathway have been partially characterized. In our study, the inactivation of a gene encoding a nonribosomal peptide synthetase in OTA producing A. carbonarius ITEM 5010 has removed the ability of the fungus to produce OTA. This is themore » first report on the involvement of an nrps gene product in OTA biosynthetic pathway in Aspergillus species. The absence of OTA and ochratoxin α-the isocoumaric derivative of OTA, and the concomitant increase of ochratoxin β- the dechloro analog of ochratoxin α- were observed in the liquid culture of transformed strain. The data provide the first evidence that the enzymatic step adding phenylalanine to polyketide dihydroisocoumarin precedes the chlorination step to form OTA in A. carbonarius, and that ochratoxin α is a product of hydrolysis of OTA, giving an interesting new insight in the biosynthetic pathway of the toxin.« less

Biological Modification of Trichothecene Mycotoxins: Acetylation and Deacetylation of Deoxynivalenols by Fusarium spp

PubMed Central

Yoshizawa, Takumi; Morooka, Nobuichi

1975-01-01

Attempts were made to elucidate the acetyl transformation of novel trichothecene mycotoxins, 3α,7α,15-trihydroxy-12,13-epoxytrichothec-9-en-8-one (deoxynivalenol) and its derivatives, by trichothecene-producing strains of Fusarium nivale, F. roseum, and F. solani. In the peptone-supplemented Czapek-Dox medium, F. roseum converted 3α-acetoxy-7α,15-dihydroxy-12,13-epoxytrichothec-9-en-8-one (3-acetyldeoxynivalenol) to deoxynivalenol. 3-Acetyldeoxynivalenol was also deacetylated by intact mycelia of the three strains in sugar-free Czapek-Dox medium. The growing F. nivale acetylated deoxynivalenol to afford a small amount of 3-acetyldeoxynivalenol. 3α,7α,15-Triacetoxy-12,13-epoxytrichothec-9-en-8-one (deoxynivalenol triacetate) was transformed by the intact mycelium of F. solani into 7α,15-diacetoxy-3α-hydroxy-12,13-epoxytrichothec-9-en-8-one (7,15-diacetyl-deoxynivalenol), which was then deacetylated to give 7α-acetoxy-3α,15-dihydroxy-12,13-epoxytrichothec-9-en-8-one (7-acetyldeoxynivalenol). It was noted that the ester at C-7 was not hydrolyzed by the fungal mycelium. PMID:234156

The origin, distribution, and evolution of Type A trichothecenes in the Fusarium graminearum species complex

USDA-ARS?s Scientific Manuscript database

Members of the Fusarium graminearum species complex (FGSC) are the major cause of Fusarium Head Blight (FHB) of cereal crops worldwide. FGSC strains typically produce one of three B trichothecenes (3ADON, 15ADON, NIV), which can contaminate grain and have toxic effects in animals and humans. Product...

Ochratoxin A Contamination of Food from Croatia

PubMed Central

Peraica, Maja; Flajs, Dubravka; Domijan, Ana-Marija; Ivić, Dario; Cvjetković, Bogdan

2010-01-01

Ochratoxin A (OTA) is a mycotoxin with nephrotoxic, genotoxic and carcinogenic properties produced by Penicillium and Aspergillus moulds under different climatic conditions. Humans and animals are exposed to this compound mainly via ingestion of contaminated food. In Croatia, research on mycotoxins focused on OTA when the mycotoxin theory of endemic nephropathy (EN) was postulated. Ochratoxin A was more frequent and at higher concentration in foods from EN than those from the control regions. Subsequently, OTA concentrations were determined in some commodities intended for human consumption such as maize, wheat, beans and wine. Samples from all parts of Croatia were analyzed and OTA was found in all types of commodities. It was frequently found together with other mycotoxins (fumonisin B1, fumonisin B2 and zearalenone). In general, OTA concentration in foods from Croatia is low, but the frequency of positive samples shows considerable variations from year to year depending also on sampling location. Although low levels of OTA were found in a large proportion of analyzed food samples, its persistent co-occurrence with other significant mycotoxins should raise serious public health concerns as there interactions may be synergistic or additive in causing toxicity in humans and animals. There is need to establish control measures through which such contaminations in foods can be managed. PMID:22069674

Genetic variation for resistance to ergot (Claviceps purpurea [Fr.] Tul.) among full-sib families of five populations of winter rye (Secale cereale L.).

PubMed

Mirdita, V; Dhillon, B S; Geiger, H H; Miedaner, T

2008-12-01

Ergot (Claviceps purpurea [Fr.] Tul.) is a serious disease of rye (Secale cereale L.) and it adversely affects the quality of grain. The present investigation was undertaken to study genotypic variability among full-sib families (FSF) of five open-pollinated (OP) winter rye populations of highly diverse origin, namely Dankovskie Selekcyine (Poland), Charkovskaja (Ukraine), NEM4 (Russia), Halo and Carokurz, both from Germany. About 50 FSF were developed at random in each population, and the FSF of each population were evaluated in separate but adjacent experiments conducted in four environments under artificial inoculation. A mixture of conidia of C. purpurea isolates was sprayed thrice during the flowering period. The materials were manually harvested at yellow-ripe stage. Resistance trait recorded was disease severity, i.e. percent ergot sclerotia in grain by weight. Mean ergot severity ranged from 2.29 to 4.08% for the five populations across environments. Significant genotypic variation (P < 0.01) due to FSF and FSF x environment interaction was observed within each population. Genotypic variation within all populations was higher than that among five populations. All populations showed high estimates of heritability (0.72-0.89). The study indicated that the evaluated OP populations are rich reservoirs of genetic variation that should also be used in hybrid breeding. Recurrent selection to further improve ergot resistance should be successful.

Ochratoxin production and taxonomy of the yellow aspergilli (Aspergillus section Circumdati)

PubMed Central

Visagie, C.M.; Varga, J.; Houbraken, J.; Meijer, M.; Kocsubé, S.; Yilmaz, N.; Fotedar, R.; Seifert, K.A.; Frisvad, J.C.; Samson, R.A.

2014-01-01

Aspergillus section Circumdati or the Aspergillus ochraceus group, includes species with rough walled stipes, biseriate conidial heads, yellow to ochre conidia and sclerotia that do not turn black. Several species are able to produce mycotoxins including ochratoxins, penicillic acids, and xanthomegnins. Some species also produce drug lead candidates such as the notoamides. A polyphasic approach was applied using morphological characters, extrolite data and partial calmodulin, β-tubulin and ITS sequences to examine the evolutionary relationships within this section. Based on this approach the section Circumdati is revised and 27 species are accepted, introducing seven new species: A. occultus, A. pallidofulvus, A. pulvericola, A. salwaensis, A. sesamicola, A. subramanianii and A. westlandensis. In addition we correctly apply the name A. fresenii (≡ A. sulphureus (nom. illeg.)). A guide for the identification of these 27 species is provided. These new species can be distinguished from others based on morphological characters, sequence data and extrolite profiles. The previously described A. onikii and A. petrakii were found to be conspecific with A. ochraceus, whilst A. flocculosus is tentatively synonymised with A. ochraceopetaliformis, despite extrolite differences between the two species. Based on the extrolite data, 13 species of section Circumdati produce large amounts of ochratoxin A: A. affinis, A. cretensis, A. fresenii, A. muricatus, A. occultus, A. ochraceopetaliformis (A. flocculosus), A. ochraceus, A. pseudoelegans, A. pulvericola, A. roseoglobulosus, A. sclerotiorum, A. steynii and A. westerdijkiae. Seven additional species produce ochratoxin A inconsistently and/or in trace amounts: A. melleus, A. ostianus, A. persii, A. salwaensis, A. sesamicola, A. subramanianii and A. westlandensis. The most important species regarding potential ochratoxin A contamination in agricultural products are A. ochraceus, A

Biomonitoring using dried blood spots: detection of ochratoxin A and its degradation product 2'R-ochratoxin A in blood from coffee drinkers.

PubMed

Cramer, Benedikt; Osteresch, Bernd; Muñoz, Katherine A; Hillmann, Hartmut; Sibrowski, Walter; Humpf, Hans-Ulrich

2015-09-01

In this study, human exposure to the mycotoxin ochratoxin A (OTA) and its thermal degradation product 2'R-ochratoxin A (2'R-OTA, previously named as 14R-Ochratoxin A [22]) through coffee consumption was assessed. LC-MS/MS and the dried blood spot (DBS) technique were used for the analysis of blood samples from coffee and noncoffee drinkers (n = 50), and food frequency questionnaires were used to document coffee consumption. For the detection of OTA and 2'R-OTA in blood, a new sensitive and efficient sample preparation method based on DBS was established and validated. Using this technique 2'R-OTA was for the first time detected in biological samples. Comparison between coffee drinkers and noncoffee drinkers showed for the first time that 2'R-OTA was only present in blood from the first group while OTA could be found in both groups in a mean concentration of 0.21 μg/L. 2'R-OTA mean concentration was 0.11 μg/L with a maximum concentration of 0.414 μg/L. Thus, in average 2'R-OTA was approx. half the concentration of OTA but in some cases even exceeded OTA levels. No correlation between the amounts of coffee consumption and OTA or 2'R-OTA levels was observed. The results of this study revealed for the first time a high exposure of coffee consumers to 2'R-OTA, a compound formed from OTA during coffee roasting. Since little information is available regarding toxicity and possible carcinogenicity of this compound, further OTA monitoring in blood including 2'R-OTA is advisable. © 2015 The Authors. Molecular Nutrition & Food Research published by Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Mycotoxin contamination of animal feedingstuff: detoxification by gamma-irradiation and reduction of aflatoxins and ochratoxin A concentrations.

PubMed

Di Stefano, Vita; Pitonzo, Rosa; Cicero, Nicola; D'Oca, Maria Cristina

2014-01-01

Mycotoxins are fungal secondary metabolites identified in many agricultural products screened for toxigenic moulds. They have been reported to be carcinogenic, teratogenic, tremorogenic, haemorrhagic and dermatitic to a wide range of organisms. With the increasing stringent regulations for mycotoxins imposed by importing countries such as those of the European Union, many cereals that are not safe for human consumption are used in formulations intended for animal feed. Gamma-rays are reported in the scientific literature to destroy ochratoxin A and aflatoxin in food crops and feed. The present study provides preliminary data for establishing the effect of dose of gamma-irradiation, ranging from 0 to 15 kGy, on aflatoxins and ochratoxin A reduction in commercial animal feed. The mycotoxin levels were determined by means of immunoaffinity clean-up (IAC) and HPLC with fluorescence detection (HPLC-FLD). The maximum reductions found at 15 kGy were 23.9%, 18.2%, 11.0%, 21.1% and 13.6% for ochratoxin A, aflatoxin B₁, aflatoxin B₂, aflatoxin G₁ and aflatoxin G₂, respectively. Results showed that the gamma-rays even at 15 kGy were not effective in the complete destruction of ochratoxin A and aflatoxins in the tested feed.

New tricks of an old enemy: isolates of F usarium graminearum produce a type A trichothecene mycotoxin

PubMed Central

Varga, Elisabeth; Wiesenberger, Gerlinde; Hametner, Christian; Ward, Todd J.; Dong, Yanhong; Schöfbeck, Denise; McCormick, Susan; Broz, Karen; Stückler, Romana; Schuhmacher, Rainer; Krska, Rudolf; Kistler, H. Corby; Adam, Gerhard

2015-01-01

Summary The ubiquitous filamentous fungus F usarium graminearum causes the important disease Fusarium head blight on various species of cereals, leading to contamination of grains with mycotoxins. In a survey of F . graminearum (sensu stricto) on wheat in North America several novel strains were isolated, which produced none of the known trichothecene mycotoxins despite causing normal disease symptoms. In rice cultures, a new trichothecene mycotoxin (named NX‐2) was characterized by liquid chromatography‐tandem mass spectrometry. Nuclear magnetic resonance measurements identified NX‐2 as 3α‐acetoxy‐7α,15‐dihydroxy‐12,13‐epoxytrichothec‐9‐ene. Compared with the well‐known 3‐acetyl‐deoxynivalenol (3‐ADON), it lacks the keto group at C‐8 and hence is a type A trichothecene. Wheat ears inoculated with the isolated strains revealed a 10‐fold higher contamination with its deacetylated form, named NX‐3, (up to 540 mg kg−1) compared with NX‐2. The toxicities of the novel mycotoxins were evaluated utilizing two in vitro translation assays and the alga C hlamydomonas reinhardtii. NX‐3 inhibits protein biosynthesis to almost the same extent as the prominent mycotoxin deoxynivalenol, while NX‐2 is far less toxic, similar to 3‐ADON. Genetic analysis revealed a different TRI 1 allele in the N‐isolates, which was verified to be responsible for the difference in hydroxylation at C‐8. PMID:25403493

Effects of Trichothecene Production on the Plant Defense Response and Fungal Physiology: Overexpression of the Trichoderma arundinaceum tri4 Gene in T. harzianum

PubMed Central

Cardoza, R. E.; McCormick, S. P.; Malmierca, M. G.; Olivera, E. R.; Alexander, N. J.; Monte, E.

2015-01-01

Trichothecenes are fungal sesquiterpenoid compounds, the majority of which have phytotoxic activity. They contaminate food and feed stocks, resulting in potential harm to animals and human beings. Trichoderma brevicompactum and T. arundinaceum produce trichodermin and harzianum A (HA), respectively, two trichothecenes that show different bioactive properties. Both compounds have remarkable antibiotic and cytotoxic activities, but in addition, trichodermin is highly phytotoxic, while HA lacks this activity when analyzed in vivo. Analysis of Fusarium trichothecene intermediates led to the conclusion that most of them, with the exception of the hydrocarbon precursor trichodiene (TD), have a detectable phytotoxic activity which is not directly related to the structural complexity of the intermediate. In the present work, the HA intermediate 12,13-epoxytrichothec-9-ene (EPT) was produced by expression of the T. arundinaceum tri4 gene in a transgenic T. harzianum strain that already produces TD after transformation with the T. arundinaceum tri5 gene. Purified EPT did not show antifungal or phytotoxic activity, while purified HA showed both antifungal and phytotoxic activities. However, the use of the transgenic T. harzianum tri4 strain induced a downregulation of defense-related genes in tomato plants and also downregulated plant genes involved in fungal root colonization. The production of EPT by the transgenic tri4 strain raised levels of erg1 expression and reduced squalene accumulation while not affecting levels of ergosterol. Together, these results indicate the complex interactions among trichothecene intermediates, fungal antagonists, and host plants. PMID:26150463

Involvement of Trichoderma trichothecenes in the biocontrol activity and in the induction of plant defense related genes

USDA-ARS?s Scientific Manuscript database

Trichoderma species produce trichothecenes, most notably trichodermin and harzianum A (HA), by a biosynthetic pathway in which several of the involved proteins have significant differences in functionality, compared to their Fusarium orthologues. In addition, the genes encoding these proteins show a...

A lipid transfer protein increases the glutathione content and enhances Arabidopsis resistance to a trichothecene mycotoxin

USDA-ARS?s Scientific Manuscript database

Fusarium head blight (FHB) or scab is one of the most important plant diseases worldwide, affecting wheat, barley and other small grains. Trichothecene mycotoxins such as deoxynivalenol (DON) accumulate in the grain, presenting a food safety risk and health hazard to humans and animals. Despite cons...

Conversion of deoxynivalenol to 3-acetyldeoxynivalenol in barley-derived fuel ethanol co-products with yeast expressing trichothecene 3-O-acetyltransferases

PubMed Central

2011-01-01

Background The trichothecene mycotoxin deoxynivalenol (DON) may be concentrated in distillers dried grains with solubles (DDGS; a co-product of fuel ethanol fermentation) when grain containing DON is used to produce fuel ethanol. Even low levels of DON (≤ 5 ppm) in DDGS sold as feed pose a significant threat to the health of monogastric animals. New and improved strategies to reduce DON in DDGS need to be developed and implemented to address this problem. Enzymes known as trichothecene 3-O-acetyltransferases convert DON to 3-acetyldeoxynivalenol (3ADON), and may reduce its toxicity in plants and animals. Results Two Fusarium trichothecene 3-O-acetyltransferases (FgTRI101 and FfTRI201) were cloned and expressed in yeast (Saccharomyces cerevisiae) during a series of small-scale ethanol fermentations using barley (Hordeum vulgare). DON was concentrated 1.6 to 8.2 times in DDGS compared with the starting ground grain. During the fermentation process, FgTRI101 converted 9.2% to 55.3% of the DON to 3ADON, resulting in DDGS with reductions in DON and increases in 3ADON in the Virginia winter barley cultivars Eve, Thoroughbred and Price, and the experimental line VA06H-25. Analysis of barley mashes prepared from the barley line VA04B-125 showed that yeast expressing FfTRI201 were more effective at acetylating DON than those expressing FgTRI101; DON conversion for FfTRI201 ranged from 26.1% to 28.3%, whereas DON conversion for FgTRI101 ranged from 18.3% to 21.8% in VA04B-125 mashes. Ethanol yields were highest with the industrial yeast strain Ethanol Red®, which also consumed galactose when present in the mash. Conclusions This study demonstrates the potential of using yeast expressing a trichothecene 3-O-acetyltransferase to modify DON during commercial fuel ethanol fermentation. PMID:21888629

Comparison of electrospray ionization and atmospheric pressure photoionization liquid chromatography mass spectrometry methods for analysis of ergot alkaloids from endophyte-infected sleepygrass (Achnatherum robustum).

PubMed

Jarmusch, Alan K; Musso, Ashleigh M; Shymanovich, Tatsiana; Jarmusch, Scott A; Weavil, Miranda J; Lovin, Mary E; Ehrmann, Brandie M; Saari, Susanna; Nichols, David E; Faeth, Stanley H; Cech, Nadja B

2016-01-05

Ergot alkaloids are mycotoxins with an array of biological effects. With this study, we investigated for the first time the application of atmospheric pressure photoionization (APPI) as an ionization method for LC-MS analysis of ergot alkaloids, and compared its performance to that of the more established technique of electrospray ionization (ESI). Samples of the grass Achnatherum robustum infected with the ergot producing Epichloë fungus were extracted using cold methanol and subjected to reserved-phase HPLC-ESI-MS and HPLC-APPI-MS analysis. The ergot alkaloids ergonovine and lysergic acid amide were detected in these samples, and quantified via external calibration. Validation parameters were recorded in accordance with ICH guidelines. A triple quadrupole MS operated in multiple reaction monitoring yielded the lowest detection limits. The performance of APPI and ESI methods was comparable. Both methods were subject to very little matrix interference, with percent recoveries ranging from 82% to 100%. As determined with HPLC-APPI-MS quantification, lysergic acid amide and ergonovine were extracted from an A. robustum sample infected with the Epichloë fungus at concentrations of 1.143±0.051 ppm and 0.2822±0.0071 ppm, respectively. There was no statistically significant difference between these concentrations and those determined using ESI for the same samples. Copyright © 2015 Elsevier B.V. All rights reserved.

Saccharomyces cerevisiae cell wall components as tools for ochratoxin a decontamination.

PubMed

Piotrowska, Małgorzata; Masek, Anna

2015-04-02

The aim of this study was to evaluate the usefulness of Saccharomyces cerevisiae cell wall preparations in the adsorption of ochratoxin A (OTA). The study involved the use of a brewer's yeast cell wall devoid of protein substances, glucans obtained by water and alkaline extraction, a glucan commercially available as a dietary supplement for animals and, additionally, dried brewer's yeast for comparison. Fourier Transform Infrared (FTIR) analysis of the obtained preparations showed bands characteristic for glucans in the resulting spectra. The yeast cell wall preparation, water-extracted glucan and the commercial glucan bound the highest amount of ochratoxin A, above 55% of the initial concentration, and the alkaline-extracted glucan adsorbed the lowest amount of this toxin. It has been shown that adsorption is most effective at a close-to-neutral pH, while being considerably limited in alkaline conditions.

Biomonitoring using dried blood spots: Detection of ochratoxin A and its degradation product 2’R‐ochratoxin A in blood from coffee drinkers*

PubMed Central

Cramer, Benedikt; Osteresch, Bernd; Muñoz, Katherine A.; Hillmann, Hartmut; Sibrowski, Walter

2015-01-01

Scope In this study, human exposure to the mycotoxin ochratoxin A (OTA) and its thermal degradation product 2’R‐ochratoxin A (2’R‐OTA, previously named as 14R‐Ochratoxin A [22]) through coffee consumption was assessed. LC‐MS/MS and the dried blood spot (DBS) technique were used for the analysis of blood samples from coffee and noncoffee drinkers (n = 50), and food frequency questionnaires were used to document coffee consumption. Methods and results For the detection of OTA and 2’R‐OTA in blood, a new sensitive and efficient sample preparation method based on DBS was established and validated. Using this technique 2’R‐OTA was for the first time detected in biological samples. Comparison between coffee drinkers and noncoffee drinkers showed for the first time that 2’R‐OTA was only present in blood from the first group while OTA could be found in both groups in a mean concentration of 0.21 μg/L. 2’R‐OTA mean concentration was 0.11 μg/L with a maximum concentration of 0.414 μg/L. Thus, in average 2’R‐OTA was approx. half the concentration of OTA but in some cases even exceeded OTA levels. No correlation between the amounts of coffee consumption and OTA or 2’R‐OTA levels was observed. Conclusion The results of this study revealed for the first time a high exposure of coffee consumers to 2’R‐OTA, a compound formed from OTA during coffee roasting. Since little information is available regarding toxicity and possible carcinogenicity of this compound, further OTA monitoring in blood including 2’R‐OTA is advisable. PMID:26012425

Effect of Neem Leaf Extract and Neem Oil on Penicillium Growth, Sporulation, Morphology and Ochratoxin A Production

PubMed Central

Mossini, Simone A. G.; Arrotéia, Carla C.; Kemmelmeier, Carlos

2009-01-01

In vitro trials were conducted to evaluate the effect of Azadirachta indica (neem) extracts on mycelial growth, sporulation, morphology and ochratoxin A production by P. verrucosum and P. brevicompactum. The effect of neem oil extract from seeds and leaf was evaluated at 0.125; 0.25 and 0.5% and 6.25 and 12.5 mg/mL, respectively, in Yeast Extract Sucrose (YES) medium. Ochratoxin A production was evaluated by a thin-layer chromatography technique. Oil extracts exhibited significant (p ≤ 0.05) reduction of growth and sporulation of the fungi. No inhibition of ochratoxin A production was observed. Given its accessibility and low cost, neem oil could be implemented as part of a sustainable integrated pest management strategy for plant disease, as it has been shown to be fungitoxic by inhibition of growth and sporulation. PMID:22069528

Effect of neem leaf extract and neem oil on Penicillium growth, sporulation, morphology and ochratoxin A production.

PubMed

Mossini, Simone A G; Arrotéia, Carla C; Kemmelmeier, Carlos

2009-09-01

In vitro trials were conducted to evaluate the effect of Azadirachtaindica (neem) extracts on mycelial growth, sporulation, morphology and ochratoxin A production by P. verrucosum and P. brevicompactum. The effect of neem oil extract from seeds and leaf was evaluated at 0.125; 0.25 and 0.5% and 6.25 and 12.5 mg/mL, respectively, in Yeast Extract Sucrose (YES) medium. Ochratoxin A production was evaluated by a thin-layer chromatography technique. Oil extracts exhibited significant (p ≤ 0.05) reduction of growth and sporulation of the fungi. No inhibition of ochratoxin A production was observed. Given its accessibility and low cost, neem oil could be implemented as part of a sustainable integrated pest management strategy for plant disease, as it has been shown to be fungitoxic by inhibition of growth and sporulation.

Impact of competitive fungi on Trichothecene production by Fusarium graminearum.

PubMed

Cooney, J M; Lauren, D R; di Menna, M E

2001-01-01

Bioassays were used to determine the production of the trichothecene mycotoxin, deoxynivalenol (DON), by two isolates of Fusarium graminearum when grown in association with potentially competitive fungi and an antifungal chemical, 6-pentyl-alpha-pyrone (6PAP). The presence of 6PAP in the culture medium reduced DON production by as much as 80%, but this effect was reduced for the F. graminearum isolate that most efficiently metabolized the added 6PAP. A 6PAP-producing Trichoderma isolate grown in a competition assay system with the F. graminearum isolates was also able to substantially reduce DON production. When Fusarium isolates (F. crookwellense, F. culmorum, F. subglutinans, F. poae, F. equiseti, F. avenaceum, and F. sambucinum), which co-occur with F. graminearum in New Zealand maize plants (Zea mays), were grown in competition assays, the effect on DON production was variable. However, all isolates of F. subglutinans tested were shown to cause reductions in DON production (by 13-76%, mean = 62%). F. subglutinans frequently co-occurs with F. graminearum, but its presence can vary with location and time of the season. When the competitive fungus tested was also a trichothecene producer (e.g., of nivalenol), both toxins were produced in the assay medium. The results indicate that mycotoxin production by F. graminearum can be affected by the presence of particular competitive fungi. These results have implications for an ecological understanding of pathogenicity and of mycotoxin accumulation in plants. Early establishment of F. subglutinans, for example, may act as a biological control mechanism providing a temporary protection against invasion by more commonly toxigenic fusaria such as F. graminearum.

Effects of Trichothecene Production on the Plant Defense Response and Fungal Physiology: Overexpression of the Trichoderma arundinaceum tri4 Gene in T. harzianum.

PubMed

Cardoza, R E; McCormick, S P; Malmierca, M G; Olivera, E R; Alexander, N J; Monte, E; Gutiérrez, S

2015-09-01

Trichothecenes are fungal sesquiterpenoid compounds, the majority of which have phytotoxic activity. They contaminate food and feed stocks, resulting in potential harm to animals and human beings. Trichoderma brevicompactum and T. arundinaceum produce trichodermin and harzianum A (HA), respectively, two trichothecenes that show different bioactive properties. Both compounds have remarkable antibiotic and cytotoxic activities, but in addition, trichodermin is highly phytotoxic, while HA lacks this activity when analyzed in vivo. Analysis of Fusarium trichothecene intermediates led to the conclusion that most of them, with the exception of the hydrocarbon precursor trichodiene (TD), have a detectable phytotoxic activity which is not directly related to the structural complexity of the intermediate. In the present work, the HA intermediate 12,13-epoxytrichothec-9-ene (EPT) was produced by expression of the T. arundinaceum tri4 gene in a transgenic T. harzianum strain that already produces TD after transformation with the T. arundinaceum tri5 gene. Purified EPT did not show antifungal or phytotoxic activity, while purified HA showed both antifungal and phytotoxic activities. However, the use of the transgenic T. harzianum tri4 strain induced a downregulation of defense-related genes in tomato plants and also downregulated plant genes involved in fungal root colonization. The production of EPT by the transgenic tri4 strain raised levels of erg1 expression and reduced squalene accumulation while not affecting levels of ergosterol. Together, these results indicate the complex interactions among trichothecene intermediates, fungal antagonists, and host plants. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Development and validation of a LC-MS method for quantitation of ergot alkaloids in lateral saphenous vein tissue

USDA-ARS?s Scientific Manuscript database

A liquid chromatography-mass spectrometry (LC/MS) method for simultaneous quantitation of seven ergot alkaloids (lysergic acid, ergonovine, ergovaline, ergocornine, ergotamine, ergocryptine and ergocrystine) in vascular tissue was developed and validated. Reverse-phase chromatography, coupled to an...

Comparison of In-Solution Biorecognition Properties of Aptamers against Ochratoxin A

PubMed Central

McKeague, Maureen; Velu, Ranganathan; De Girolamo, Annalisa; Valenzano, Stefania; Pascale, Michelangelo; Smith, McKenzie; DeRosa, Maria C.

2016-01-01

Ochratoxin A (OTA) is a mycotoxin produced as a secondary metabolite by several species of Aspergillus and Penicillium and frequently found as a natural contaminant in a wide range of food commodities. Novel and robust biorecognition agents for detecting this molecule are required. Aptamers are artificial nucleic acid ligands able to bind with high affinity and specificity to a given target molecule. In the last few years, three separate research groups have selected aptamers for ochratoxin A. While each of these three families of aptamers have been incorporated into various methods for detecting OTA, it is unclear if each aptamer candidate is better suited for a particular application. Here, we perform the first head-to-head comparison of solution-based binding parameters for these groups of aptamers. Based on our results, we provide recommendations for the appropriate choice of aptamer for incorporation into solution-based biorecognition assays and applications. PMID:27854269

Saccharomyces Cerevisiae Cell Wall Components as Tools for Ochratoxin A Decontamination

PubMed Central

Piotrowska, Małgorzata; Masek, Anna

2015-01-01

The aim of this study was to evaluate the usefulness of Saccharomyces cerevisiae cell wall preparations in the adsorption of ochratoxin A (OTA). The study involved the use of a brewer’s yeast cell wall devoid of protein substances, glucans obtained by water and alkaline extraction, a glucan commercially available as a dietary supplement for animals and, additionally, dried brewer’s yeast for comparison. Fourier Transform Infrared (FTIR) analysis of the obtained preparations showed bands characteristic for glucans in the resulting spectra. The yeast cell wall preparation, water-extracted glucan and the commercial glucan bound the highest amount of ochratoxin A, above 55% of the initial concentration, and the alkaline-extracted glucan adsorbed the lowest amount of this toxin. It has been shown that adsorption is most effective at a close-to-neutral pH, while being considerably limited in alkaline conditions. PMID:25848694

A risk assessment of dietary ochratoxin A in the United States

USDA-ARS?s Scientific Manuscript database

Background: Ochratoxin A (OTA) is a mycotoxin (fungal toxin) found in multiple foodstuffs. Because OTA has been shown to cause kidney disease in multiple species, several governmental bodies around the world have set maximum allowable levels of OTA in different foods and beverages. Objectives: In ...

Macrocyclic trichothecenes are undetectable in kudzu (Pueraria montana) plants treated with a high-producing isolate of Myrothecium verrucaria.

PubMed

Abbas, H K; Tak, H; Boyette, C D; Shier, W T; Jarvis, B B

2001-09-01

Myrothecium verrucaria was found to be an effective pathogen against kudzu grown in the greenhouse and the field. M. verrucaria produced large amounts of macrocyclic trichothecenes when cultured on solid rice medium, including epiroridin E (16.8 mg/g crude extract), epiisororidin E (1 mg/g), roridin E (8.7 mg/g), roridin H (31.3 mg/g), trichoverrin A (0.6 mg/g), trichoverrin B (0.1 mg/g), verrucarin A (37.4 mg/g), and verrucarin J (2.2 mg/g). Most of these toxins were also isolated from M. verrucaria spores and mycelia grown on potato dextrose agar medium, including epiroridin E (32.3 mg/g), epiisororidin E (28.6 mg/g), roridin E (0 mg/g), roridin H (60 mg/g), trichoverrin A (1.3 mg/g), trichoverrin B (1.8 mg/g), verrucarin A (13.8 mg/g), and verrucarin J (131 mg/g). When M. verrucaria was cultured on liquid media, the numbers but not the amounts of toxins decreased. Only epiroridin E (28.3 mg/g), epiisororidin E (29.6 mg/g), verrucarin B (195 mg/g) and verrucarin J (52.6 mg/g) were measured when the fungus was cultured on cornsteep medium. On soyflour-cornmeal broth M. verrucaria produced several toxins, including epiroridin E (58.1 mg/g), epiisororidin E (5.8 mg/g), verrucarin B (29.9 mg/g) and verrucarin J (32 mg/g). In contrast, no macrocyclic trichothecenes were detected by HPLC analysis of plant tissues of kudzu, sicklepod, and soybean treated with aqueous suspensions of M. verrucaria spores formulated with a surfactant. Chloroform-methanol extracts of kudzu leaves and stems treated with M. verrucaria spores were less cytotoxic to four cultured mammalian cell lines than the corresponding extracts from control plants. Purified macrocyclic trichothecenes (verrucarin A and T-2 toxin) were very cytotoxic to the same cell lines (< or = 2 ng/ml). These results show that neither intact macrocyclic trichothecenes nor toxic metabolites could be detected in plant tissues after treatment with M. verrucaria spores. These results argue for both safety and efficacy for the

Vasoconstrictive Responses by the Carotid and Auricular Arteries in goats to Ergot Alkaloid Exposure

NASA Astrophysics Data System (ADS)

Aiken, Glen; Flythe, Michael

2014-11-01

A fungal endophyte (Neotyphodium coenophialum) infects most plants of ‘Kentucky 31’ tall fescue (Lolium arundinaceum) and produces ergot alkaloids that cause persistent constriction of the vascular system in grazing livestock. Consequently, animals undergoing this toxicosis cannot regulate core body temperature and are vulnerable to heat and cold stresses. An experiment was conducted to determine if the caudal and auricular arteries in goats (Capra aegagrus hircus) vasoconstrict in response to ergot alkaloids. Seven, rumen fistulated goats were fed ad libitum orchardgrass (Dactylis glomeratia) hay and ruminally infused with endophtye-free seed (E-) for a 7-day adjustment period. Two periods followed with E- and endophyte-infected (E+) seed being randomly assigned to the 2 goat groups in period 1 and then switching treatments between groups in period 2. Infused E+ and E- seed were in equal proportions to the hay such that concentrations of ergovaline and ergovalanine were 0.80 µg per g dry matter for the E+ treatment. Cross-sections of both arteries were imaged using Doppler ultrasonography on days 0, 2, 4, 6, 8, and 12 in period 1 and on days 0, 1, 2, 3, 6, 7, and 9 in period 2. Differences from average baseline areas were used to determine presence or absence of alkaloid-induced vasoconstriction. Carotid arteries initiated constriction on imaging day 2 in both periods, and auricular arteries initiated constriction on imaging day 2 in period 1 and on day 6 in period 2. Luminal areas of the carotid arteries in E+ goats were 46% less than baseline areas in both periods after vasoconstriction occurred, whereas auricular arteries in E+ goats were 52% less than baseline areas in period 1 and 38% in period 2. Both arteries in E+ goats in period 1 relaxed relative to baseline areas by imaging day 2 after they were switched to the E- treatment. Results indicated that goats can vasoconstrict when exposed to ergot alkaloids that could disrupt their thermoregulation.

Monitoring of aflatoxins and ochratoxin A in Czechoslovak human sera by immunoassay

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fukal, L.; Reisnerova, H.

1990-03-01

Since a level of food contamination with aflatoxins and ochratoxin A has been found low in Czechoslovakia, human exposure to these mycotoxins may not be negligible. However, analysis of food samples provides only indirect evidence of mycotoxin ingestion and no evidence about mycotoxin absorption. Direct evidence can only be obtained by analysis of human body fluids. Therefore, the authors decided to carry out a monitoring of aflatoxin and ochratoxin A level in human sera. In general, TLC and HPLC are most commonly used to analyze mycotoxins and its metabolites. The recent development of immunochemical techniques opens the possibility of determiningmore » individual exposure in a relatively large human population. These assays have the advantage of high specificity and sensitivity. Sample through-put is high, and the methods are technically simple and can be performed at low cost.« less

Occurrence of ochratoxin A-producing fungi in raw Brazilian coffee.

PubMed

Urbano, G R; Taniwaki, M H; Leitão, M F; Vicentini, M C

2001-08-01

Ochratoxin A (OA)-producing fungi were identified in coffee at different stages of maturation. The toxin was quantified in coffee during terrace drying and in coffee stored in barns. By direct plating, a high level of contamination (100%) was found in the coffee beans studied, with the genus Aspergillus representing 33.2%, of which Aspergillus ochraceus and Aspergillus niger represented 10.3 and 22.9%, respectively, of the strains isolated from the coffee beans. The capacity to produce ochratoxin was determined in 155 strains of A. ochraceus and A. niger using both the agar plug method and extraction with chloroform, giving positive results for 88.1% of the A. ochraceus strains and 11.5% of the A. niger strains. Analysis for OA in the terrace and barn coffee samples showed that, independent of cultivar, year harvested, or production region, all except one of the samples analyzed showed mycotoxin levels below the limit suggested by the European Common Market (8 microg/kg), thus indicating that the problem is restricted and due to severe faults in harvesting and storage practices.

Gut satiety hormones cholecystokinin and glucagon-like Peptide-17-36 amide mediate anorexia induction by trichothecenes T-2 toxin, HT-2 toxin, diacetoxyscirpenol and neosolaniol.

PubMed

Zhang, Jie; Liu, Shengli; Zhang, Hua; Li, Yuanyuan; Wu, Wenda; Zhang, Haibin

2017-11-15

The food-borne trichothecene mycotoxins have been documented to cause human and animal food poisoning. Anorexia is a hallmark of the trichothecene mycotoxins-induced adverse effects. Type B trichothecenes have been previously demonstrated to elicit robust anorectic responses, and this response has been directly linked to secretion of the gut satiety hormones cholecystokinin (CCK) and glucagon-like peptide-1 7-36 amide (GLP-1). However, less is known about the anorectic effects and underlying mechanisms of the type A trichothecenes, including T-2 toxin (T-2), HT-2 toxin (HT-2), diacetoxyscirpenol (DAS), neosolaniol (NEO). The purpose of this study was to relate type A trichothecenes T-2, HT-2, DAS and NEO-induced anorectic response to changes plasma concentrations of CCK and GLP-1. Following both oral gavage and intraperitoneal (IP) administration of 1mg/kg bw T-2, HT-2, DAS and NEO evoked robust anorectic response and secretion of CCK and GLP-1. Elevations of plasma CCK markedly corresponded to anorexia induction by T-2, HT-2, DAS and NEO. Following oral exposure, plasma CCK was peaked at 6h, 6h, 2h, 2h and lasted up to 24h, 24h, > 6h, > 6h for T-2, HT-2, DAS and NEO, respectively. IP exposed to four toxins all induced elevation of CCK with peak point and duration at 6h and >24h, respectively. In contrast to CCK, GLP-1 was moderately elevated by these toxins. Following both oral and IP exposure, T-2 and HT-2 evoked plasma GLP-1 elevation with peak point and duration at 2h and 6h, respectively. Plasma GLP-1 was peaked at 2h and still increased at 6h for IP and oral administration with DAS and NEO, respectively. In conclusion, CCK plays a contributory role in anorexia induction but GLP-1 might play a lesser role in this response. Copyright © 2017 Elsevier Inc. All rights reserved.

A Polyketide Synthase Encoded by the Gene An15g07920 Is Involved in the Biosynthesis of Ochratoxin A in Aspergillus niger.

PubMed

Zhang, Jian; Zhu, Liuyang; Chen, Haoyu; Li, Min; Zhu, Xiaojuan; Gao, Qiang; Wang, Depei; Zhang, Ying

2016-12-28

The polyketide synthase gene An15g07920 was known in Aspergillus niger CBS 513.88 as putatively involved in the production of ochratoxin A (OTA). Genome resequencing analysis revealed that the gene An15g07920 is also present in the ochratoxin-producing A. niger strain 1062. Disruption of An15g07920 in A. niger 1062 removed its capacity to biosynthesize ochratoxin β (OTβ), ochratoxin α (OTα), and OTA. These results indicate that the polyketide synthase encoded by An15g07920 is a crucial player in the biosynthesis of OTA, in the pathway prior to the phenylalanine ligation step. The gene An15g07920 reached its maximum transcription level before OTA accumulation reached its highest level, confirming that gene transcription precedes OTA production. These findings will not only help explain the mechanism of OTA production in A. niger but also provide necessary information for the development of effective diagnostic, preventive, and control strategies to reduce the risk of OTA contamination in foods.

Diversity of Fusarium head blight populations and trichothecene toxin types reveals regional differences in pathogen composition and temporal dynamics

USDA-ARS?s Scientific Manuscript database

Analyses of genetic diversity, trichothecene genotype composition, and population structure were conducted using 4,086 Fusarium graminearum isolates collected from wheat in eight Canadian provinces over a three year period between 2005 and 2007. The results revealed substantial regional differences ...

The effect of pomelo citrus (Citrus maxima var. Nambangan), vitamin C and lycopene towards the number reduction of mice (Mus musculus) apoptotic hepatocyte caused of ochratoxin A

NASA Astrophysics Data System (ADS)

Badriyah, Hastuti, Utami Sri

2017-06-01

Foods can contaminated by some mycotoxin produced by molds. Ochratoxin A is a sort of mycotoxin that cause structural damage on hepatocytes. Pomelo citrus (Citrus maxima var. Nambangan) contain vitamin C and lycopene that have antioxidant character. This research is done to: 1)examine the effect of pomelo citrus juice, vitamin C, and lycopene treatment towards the number reduction of mice apoptotic hepatocytes caused by ochratoxin A exposure, 2)examine the effect of vitamin C mixed with lycopene treatment towards the number reduction of mice apoptotic hepatocytes caused by ochratoxin A exposure. The experimental group used male mice strain BALB-C in the age of three month and bodyweight 20-30 grams devided in 4 experiment group and control group. The experiment group I were administered pomelo citrus juice 0,5 ml/30 grams BW/day orally during 2 weeks and then administered with ochratoxin in the dose of 1 mg/kg BW during 1 week. The experiment group II were administered with vitamin C in the dose of 5,85 µg/30g BW with the same methods. The experiment group III were administered with lycopene in the dose of 0,1025 µg/30 g BW with the same methods. The experiment group IV were administered with vitamin C mixed with lycopene with the same methods. The control group were administered with ochratoxin A in the dose of 1 mg/kg BW per oral during 1 week. The apoptotic hepatocyte number were count by microscopic observation of hepatocyte slides from experiment group as well as control group with cytochemical staining. The research result shows that: 1) the pomelo citrus juice, vitamin C as well as lycopene administration could reduce the mice apoptotic hepatocyte number caused by ochratoxin A exposure, compared with the mice apoptotic hepatocyte number caused by ochratoxin A exposure only; 2) the vitamin C mixed with lycopene could reduce the mice apoptotic hepatocyte number caused by ochratoxin A exposure compared with the mice apoptotic hepatocyte number caused by

Sorption of Ochratoxin A from aqueous solutions using beta-cyclodextrin-polyurethane polymer

USDA-ARS?s Scientific Manuscript database

The ability of a cyclodextrin-polyurethane polymer to remove ochratoxin A from aqueous solutions, including wine, was examined by batch rebinding assays and equilibrium sorption isotherms. The results were fit to two parameter models. Freundlich analysis of the sorption isotherm indicates the polyme...

Identification of a trichothecene gene cluster and description of the harzianum A biosynthesis pathway in the fungus Trichoderma arundinaceum

USDA-ARS?s Scientific Manuscript database

Trichothecenes are sesquiterpenes that act like mycotoxins. Their biosynthesis has been mainly studied in the fungal genera Fusarium, where most of the biosynthetic genes (tri) are grouped in a cluster regulated by ambient conditions and regulatory genes. Unexpectedly, few studies are available abou...

Production of Penicillic Acid and Ochratoxin A on Poultry Feed by Aspergillus ochraceus: Temperature and Moisture Requirements

PubMed Central

Bacon, C. W.; Sweeney, J. G.; Robbins, J. D.; Burdick, D.

1973-01-01

A strain of Aspergillus ochraceus Wilhelm, isolated from poultry feed, produced both penicillic acid and ochratoxin A. Studies demonstrating the ability of this fungus to colonize poultry feed and produce these two mycotoxins under various temperatures and moistures indicated that the interaction was complex. The optimal temperature for conidial development did not vary with moisture, but accumulation of both toxins did. A combination of low temperature, 15 or 22 C, and low moisture favored the production of penicillic acid, whereas high temperature, 30 C, and high moisture favored the production of ochratoxin A. PMID:4795527

Tall fescue seed extraction and partial purification of ergot alkaloids

PubMed Central

Ji, Huihua; Fannin, F.; Klotz, J.; Bush, Lowell

2014-01-01

Many substances in the tall fescue/endophyte association (Schedonorus arundinaceus/Epichloë coenophiala) have biological activity. Of these compounds only the ergot alkaloids are known to have significant mammalian toxicity and the predominant ergot alkaloids are ergovaline and ergovalinine. Because synthetically produced ergovaline is difficult to obtain, we developed a seed extraction and partial purification protocol for ergovaline/ergovalinine that provided a biologically active product. Tall fescue seed was ground and packed into several different sized columns for liquid extraction. Smaller particle size and increased extraction time increased efficiency of extraction. Our largest column was a 114 × 52 × 61 cm (W × L × D) stainless steel tub. Approximately 150 kg of seed could be extracted in this tub. The extraction was done with 80% ethanol. When the solvent front migrated to bottom of the column, flow was stopped and seed was allowed to steep for at least 48 h. Light was excluded from the solvent from the beginning of this step to the end of the purification process. Following elution, ethanol was removed from the eluate by evaporation at room temperature and the resulting syrup was freeze-dried. About 80% recovery of alkaloids was achieved with 18-fold increase in concentration of ergovaline. Initial purification of the dried product was accomplished by extracting with hexane/water (6:1, v/v). The aqueous fraction was extracted with chloroform, the aqueous layer discarded, after which the chloroform was removed with a resulting 20-fold increase of ergovaline. About 65% of the ergovaline was recovered from the chloroform residue for an overall recovery of 50%. The resultant partially purified ergovaline had biological activities in in vivo and in vitro bovine bioassays that approximate that of synthetic ergovaline. PMID:25566528

Tall fescue seed extraction and partial purification of ergot alkaloids

NASA Astrophysics Data System (ADS)

Bush, Lowell

2014-12-01

Many substances in the tall fescue/endophyte association (Schedonorus arundinaceus/Epichloë coenophiala) have biological activity. Of these compounds only the ergot alkaloids are known to have significant mammalian toxicity and the predominant ergot alkaloids are ergovaline and ergovalinine. Because synthetically produced ergovaline is difficult to obtain, we developed a seed extraction and partial purification protocol for ergovaline/ergovalinine that provided a biologically active product. Tall fescue seed was ground and packed into several different sized columns for liquid extraction. Smaller particle size and increased extraction time increased efficiency of extraction. Our largest column was a 114 × 52 × 61 cm (W×L×D) stainless steel tub. Approximately 150 kg of seed could be extracted in this tub. The extraction was done with 80% ethanol. When the solvent front migrated to bottom of the column, flow was stopped and seed was allowed to steep for at least 48 h. Light was excluded from the solvent from the beginning of this step to the end of the purification process. Following elution, ethanol was removed from the eluate by evaporation at room temperature. Resulting syrup was freeze-dried. About 80% recovery of alkaloids was achieved with 18-fold increase in concentration of ergovaline. Initial purification of the dried product was accomplished by extracting with hexane/water (6:1, v/v) and the hexane fraction was discarded. The aqueous fraction was extracted with chloroform, the aqueous layer discarded, after which the chloroform was removed with a resulting 20-fold increase of ergovaline. About 65% of the ergovaline was recovered from the chloroform residue for an overall recovery of 50%. The resultant partially purified ergovaline had biological activities in in vivo and in vitro bovine bioassays that approximate that of synthetic ergovaline.

Screening of Balansia epichloe-infected grass species for in situ ergot alkaloids using laser ablation electrospray ionization-mass spectrometry

USDA-ARS?s Scientific Manuscript database

The Balansia are clavicipitaceous symbiotic species associated with various species of tropical grasses. Laboratory culture procedures established that the Balansia species are often conspecific with grasses in tall fescue pastures that produced ergot alkaloids. However, any effects of hosts on the...

Multiplex polymerase chain reaction assay for the differential detection of trichothecene- and fumonisin-producing species of Fusarium in cornmeal.

PubMed

Bluhm, B H; Flaherty, J E; Cousin, M A; Woloshuk, C P

2002-12-01

The genus Fusarium comprises a diverse group of fungi including several species that produce mycotoxins in food commodities. In this study, a multiplex polymerase chain reaction (PCR) assay was developed for the group-specific detection of fumonisin-producing and trichothecene-producing species of Fusarium. Primers for genus-level recognition of Fusarium spp. were designed from the internal transcribed spacer regions (ITS1 and ITS2) of rDNA. Primers for group-specific detection were designed from the TRI6 gene involved in trichothecene biosynthesis and the FUM5 gene involved in fumonisin biosynthesis. Primer specificity was determined by testing for cross-reactivity against purified genomic DNA from 43 fungal species representing 14 genera, including 9 Aspergillus spp., 9 Fusarium spp., and 10 Penicillium spp. With purified genomic DNA as a template, genus-specific recognition was observed at 10 pg per reaction; group-specific recognition occurred at 100 pg of template per reaction for the trichothecene producer Fusarium graminearum and at 1 ng of template per reaction for the fumonisin producer Fusarium verticillioides. For the application of the PCR assay, a protocol was developed to isolate fungal DNA from cornmeal. The detection of F. graminearum and its differentiation from F. verticillioides were accomplished prior to visible fungal growth at <10(5) CFU/g of cornmeal. This level of detection is comparable to those of other methods such as enzyme-linked immunosorbent assay, and the assay described here can be used in the food industry's effort to monitor quality and safety.

A new validated HPLC-FLD method for detecting ochratoxin A in dry-cured meat and in blue cheese.

PubMed

Dall'asta, C; Galaverna, G; De Dea Lindner, J; Virgili, R; Neviani, E; Dossena, A

2007-09-01

In the present study, a fast and sensitive method for the quantification of ochratoxin A in two lipidicproteic food matrices has been developed. In particular, the sample preparation procedure has been optimized for dry-cured meat products and blue cheeses and tested for several validation parameters (LOD, LOQ, recovery, repeatability and within-laboratory precision). The procedure has been then applied to several dry-cured meat products and blue cheeses from the market.Ochratoxin A has been occasionally found in dry-cured and smoked ham from the market and the contamination occurred both in the outer and in the inner part of the products. Concerning the blue cheese, the occurrence of ochratoxin A is reported for the first time: OTA was occasionally found at low levels (0.1-3 μg/kg) in commercial samples of Roquefort from France and Gorgonzola from Italy, opening a new issue for risk assessment and quality control.

Determination of Aflatoxins and Ochratoxin A in Traditional Turkish Concentrated Fruit Juice Products by Multi-Immunoaffinity Column Cleanup and LC Fluorescence Detection: Single-Laboratory Validation.

PubMed

Kaymak, Tugrul; Türker, Levent; Tulay, Hüseyin; Stroka, Joerg

2018-04-27

Background : Pekmez and pestil are traditional Turkish foods made from concentrated grapejuice, which can be contaminated with mycotoxins such as aflatoxins and ochratoxin A (OTA). Objective : To carry out a single-laboratory validation of a method to simultaneously determine aflatoxins B 1 , B₂, G 1 , and G₂ and ochratoxin A in pekmez and pestil. Methods : The homogenized sample is extracted with methanol-water (80 + 20) using a high-speed blender. The (sample) extract is filtered, diluted with phosphate-buffered saline solution, and applied to a multi-immunoaffinity column (AFLAOCHRA PREP®). Aflatoxins and ochratoxin A are removed with (neat) methanol and then directly analyzed by reversed-phase LC with fluorescence detection using post-column bromination (Kobra cell®). Results : Test portions of blank pekmez and pestil were spiked with a mixture of aflatoxins and ochratoxin A to give levels ranging from 2.6 to 10.4 μg/kg and 1.0-4.0 μg/kg, respectively. Recoveries for total aflatoxins and ochratoxin A ranged from 84 to 106% and 80-97%, respectively, for spiked samples. Based on results for spiked pekmez and pestil (30 replicates each at three levels), the repeatability RSD ranged from 1.6 to 12% and 2.7-11% for total aflatoxins and ochratoxin A, respectively. Conclusions : The method performance in terms of recovery, repeatability, and detection limits has been demonstrated to be suitable for use as an Official Method. Highlights : First immunoaffinity column method validated for simultaneous analysis of aflatoxins and ochratoxin A in pekmez and pestil. Suitability for use for official purposes in Turkey, demonstrated by single-laboratory validation. Co-occurrence of aflatoxins and OTA in mulberry and carob pekmez reported for the first time.

Detection of fumonisin b1 and ochratoxin a in grain products using microsphere-based fluid array immunoassays.

PubMed

Anderson, George P; Kowtha, Vasudha A; Taitt, Chris R

2010-02-01

Grain products are a staple of diets worldwide and therefore, the ability to accurately and efficiently detect foodborne contaminants such as mycotoxins is of importance to everyone. Here we describe an indirect competitive fluid array fluoroimmunoassay to quantify the mycotoxins, fumonisin B1 and ochratoxin A. Both toxins were immobilized to the surface of microspheres using a variety of intermediate molecules and binding of biotinylated "tracer" antibody tracers determined through flow cytometry using streptavidin-phycoerythrin conjugates and the Luminex100 flow cytometer. Competitive assays were developed where the binding of biotinylated monoclonal antibodies to fumonisin B and ochratoxin A was competitively inhibited by different concentrations of those toxins in solution. Concentrations of fumonisin giving 50% inhibition were 300 pg/mL in buffer, 100 ng/g in spiked oats, and 1 μg/g in spiked cornmeal; analogous concentrations for ochratoxin A were 30 ng/mL in buffer, 30 ng/g in spiked oats, and 10 ng/g in spiked corn. The future challenge will be to expand the number of mycotoxins tested both individually and in multiplexed format using this platform.

Interaction of Ochratoxin A and Its Thermal Degradation Product 2'R-Ochratoxin A with Human Serum Albumin.

PubMed

Sueck, Franziska; Poór, Miklós; Faisal, Zelma; Gertzen, Christoph G W; Cramer, Benedikt; Lemli, Beáta; Kunsági-Máté, Sándor; Gohlke, Holger; Humpf, Hans-Ulrich

2018-06-22

Ochratoxin A (OTA) is a toxic secondary metabolite produced by several fungal species of the genus Penicillium and Aspergillus . 2′ R -Ochratoxin A (2′ R -OTA) is a thermal isomerization product of OTA formed during food processing at high temperatures. Both compounds are detectable in human blood in concentrations between 0.02 and 0.41 µg/L with 2′ R -OTA being only detectable in the blood of coffee drinkers. Humans have approximately a fifty-fold higher exposure through food consumption to OTA than to 2′ R -OTA. In human blood, however, the differences between the concentrations of the two compounds is, on average, only a factor of two. To understand these unexpectedly high 2′ R -OTA concentrations found in human blood, the affinity of this compound to the most abundant protein in human blood the human serum albumin (HSA) was studied and compared to that of OTA, which has a well-known high binding affinity. Using fluorescence spectroscopy, equilibrium dialysis, circular dichroism (CD), high performance affinity chromatography (HPAC), and molecular modelling experiments, the affinities of OTA and 2′ R -OTA to HSA were determined and compared with each other. For the affinity of HSA towards OTA, a log K of 7.0⁻7.6 was calculated, while for its thermally produced isomer 2′ R -OTA, a lower, but still high, log K of 6.2⁻6.4 was determined. The data of all experiments showed consistently that OTA has a higher affinity to HSA than 2′ R -OTA. Thus, differences in the affinity to HSA cannot explain the relatively high levels of 2′ R -OTA found in human blood samples.

Implementation of electrochemical elements for an alternative detection of ochratoxin A

NASA Astrophysics Data System (ADS)

Aristizabal, D. H.; Giraldo, D. A.; Sanchez, S.; Taborda, G.; Baeza, A.

2017-01-01

Ochratoxin A (OTA) is a nephrotoxic metabolite, hepatotoxic and carcinogenic produced mainly by Aspergillus and Penicillium fungi. Usually, the mycotoxin analysed through the technique of high performance liquid chromatography. This method is expensive and takes a lot of time. Therefore, expected to automate a device of low cost, minimal instrumentation micropolarographic (MIMP) for the analysis. For this purpose, the right programming environment and the manufacturing of the software for the graphical user interface selected automation of the MIMP through the design and simulation of the circuit, imprint and assembled in a container with connection ports. Lastly, validation and analysis of Ochratoxin A by cyclic voltammetry from analogous MIMP, automatic MIMP and a potentiostat, in order to corroborate the registered data. This way, voltammetry analysis of coumarin is obtained and the simulation of the MIMP electric circuit. Therefore, this are the key data during the investigation, because the molecule of OTA can be monitored through an acid hydrolysis of amides, composed by a phenylalanine and coumaric acid group, generating the significant decrease in the costs and time of analysis.

Oxidation of Two Hydroxylated Ochratoxin A Metabolites by Alcohol Dehydrogenase

PubMed Central

Syvertsen, Christian; Størmer, Fredrik C.

1983-01-01

(4R)-4-hydroxyochratoxin A, (4S)-4-hydroxyochratoxin A, and 10-hydroxyochratoxin A, all formed from ochratoxin A, were incubated with alcohol dehydrogenase in the presence of NAD. Only (4R)-4-hydroxyochratoxin A and 10-hydroxyochratoxin A acted as substrates for the enzyme. Km and turnover number for 10-hydroxyochratoxin A were 110 μM and 0.1 s−1, respectively. PMID:6347065

Effects of ergot alkaloid exposure on serotonin receptor mRNA in the smooth muscle of the bovine gastrointestinal tract

USDA-ARS?s Scientific Manuscript database

Various serotonin (5HT) receptor subtypes have been located in the gastrointestinal tract and some are associated with gut motility. Cattle exposed to ergot alkaloids through consumption of contaminated feedstuffs have demonstrated signs (e.g. - increased rumen DM content and total content) that sug...

Prospecting for the incidence of genes involved in ochratoxin and fumonisin biosynthesis in Brazilian strains of Aspergillus niger and Aspergillus welwitschiae.

PubMed

Massi, Fernanda Pelisson; Sartori, Daniele; de Souza Ferranti, Larissa; Iamanaka, Beatriz Thie; Taniwaki, Marta Hiromi; Vieira, Maria Lucia Carneiro; Fungaro, Maria Helena Pelegrinelli

2016-03-16

Aspergillus niger "aggregate" is an informal taxonomic rank that represents a group of species from the section Nigri. Among A. niger "aggregate" species Aspergillus niger sensu stricto and its cryptic species Aspergillus welwitschiae (=Aspergillus awamori sensu Perrone) are proven as ochratoxin A and fumonisin B2 producing species. A. niger has been frequently found in tropical and subtropical foods. A. welwitschiae is a new species, which was recently dismembered from the A. niger taxon. These species are morphologically very similar and molecular data are indispensable for their identification. A total of 175 Brazilian isolates previously identified as A. niger collected from dried fruits, Brazil nuts, coffee beans, grapes, cocoa and onions were investigated in this study. Based on partial calmodulin gene sequences about one-half of our isolates were identified as A. welwitschiae. This new species was the predominant species in onions analyzed in Brazil. A. niger and A. welwitschiae differ in their ability to produce ochratoxin A and fumonisin B2. Among A. niger isolates, approximately 32% were OTA producers, but in contrast only 1% of the A. welwitschiae isolates revealed the ability to produce ochratoxin A. Regarding fumonisin B2 production, there was a higher frequency of FB2 producing isolates in A. niger (74%) compared to A. welwitschiae (34%). Because not all A. niger and A. welwitschiae strains produce ochratoxin A and fumonisin B2, in this study a multiplex PCR was developed for detecting the presence of essential genes involved in ochratoxin (polyketide synthase and radHflavin-dependent halogenase) and fumonisin (α-oxoamine synthase) biosynthesis in the genome of A. niger and A. welwitschiae isolates. The frequency of strains harboring the mycotoxin genes was markedly different between A. niger and A. welwitschiae. All OTA producing isolates of A. niger and A. welwitschiae showed in their genome the pks and radH genes, and 95.2% of the nonproducing

Claviceps cyperi, a new cause of severe ergotism in dairy cattle consuming maize silage and teff hay contaminated with ergotised Cyperus esculentus (nut sedge) on the Highveld of South Africa.

PubMed

Naudè, T W; Botha, C J; Vorster, J H; Roux, C; Van der Linde, E J; Van der Walt, S I; Rottinghaus, G E; Van Jaarsveld, L; Lawrence, A N

2005-03-01

During December/January 1996/97 typical summer syndrome (hyperthermia and a 30% drop in milk yield) occurred in succession in two Holstein dairy herds (n=240 and n=150 milking cows, respectively) on the South African Highveld. These farms are situated in the midst of the prime maize and dairy farming areas of South Africa where this condition had never been diagnosed before. The individual components of the concentrate on both farms were negative for ergot alkaloids. Endophytic fungi and/or ergot infestation of teff and other grasses fed to the cows were then suspected of being involved, but neither endophytes nor ergot alkaloids could be implicated from these sources. By measuring the serum prolactin levels of groups of sheep (n=5) fed the first farm's total mixed ration (TMR) or its three individual fibre components for a period of 11 days, the source of the ergot alkaloids was identified. A statistically significant decrease in the level of this hormone occurred only in the group on maize silage (which constituted 28% on dry matter base of the TMR). The involvement of the maize silage was further chemically confirmed by the high levels of total ergot alkaloids, predominantly ergocryptine, found by LC-MS in the silage as well as in the TMR (115-975 ppb and 65-300 ppb, respectively). The ergot alkaloid content (mainly ergocryptine) of the maize silage on the second affected farm was 875 ppb. Withdrawal of contaminated silage resulted in gradual recovery of stock on both farms. Nut sedge (Cyperus esculentus and Cyperus rotundus of the family Cyperaceae) has a world-wide distribution and is a common weed in annual crops, and can be parasitized by Claviceps cyperi. Careful examination of the maize silage from both farms revealed that it was heavily contaminated with nut sedge and that it contained minute sclerotia, identified as those of Claviceps cyperi, originating from the latter. Nut sedge was abundant on both farms and it is believed that late seasonal rain had

Association of insects and ergot (Claviceps purpurea) in Kentucky bluegrass seed production fields.

PubMed

Butler, M D; Alderman, S C; Hammond, P C; Berry, R E

2001-12-01

Insects in Kentucky bluegrass seed production fields in Oregon, Idaho, and Washington were sampled just before harvest and their association with ergot conidia of Claviceps purpurea Fr. (Tul.) was evaluated during 1996-1998. A diversity of insects was observed at all three locations. The most abundant beneficial insects collected with sweep nets were Nysium spp., Nabis spp., ichneumonid wasps, and Hippodamia spp. The cranberry girdler, Chrysoteuchia topiaria (Zeller), was the only important pest on grass seed collected by sweep net. Numbers of aphids such as Sitobion avenae (F.), cicadellids and thrips such as Anaphothrips spp. and Aptinothrips spp. that were collected with all aphid sampler were below economic thresholds. Other insect groups occurred in low numbers. Noctuid moths collected in universal blacklight traps included nine species of cutworms and armyworms. Protogrotis obscura (B. & McD.) was the most common cutworm species and was present in all fields. The moth Chortodes rufostrigata (Pack.) previously reported only from wet meadows in northeast and south central Oregon was found in Kentucky bluegrass fields in central Oregon, suggesting that irrigated Kentucky bluegrass seed production fields may simulate a montane meadow habitat. Conidia of C. purpurea were found on a diversity of insects, including moths, flies, leafhoppers, and thrips. Up to 100% of moths and 75% of flies collected from some fields carried conidia of C. purpurea. No correlation between ergot honeydew present in a field and number of insects with conidia of C. purpurea was detected.

Development of a methodology to measure the effect of ergot alkaloids on forestomach motility using real-time wireless telemetry

USDA-ARS?s Scientific Manuscript database

The objectives of these experiments were to characterize rumen motility patterns of cattle fed once daily using a real-time wireless telemetry system, determine when to measure rumen motility with this system, and determine the effect of ruminal dosing of ergot alkaloids on rumen motility. Ruminally...

Effects of feeding bentonite clay upon ochratoxin A-induced immunosuppression in broiler chicks.

PubMed

Khatoon, Aisha; Khan, Muhammad Zargham; Abidin, Zain Ul; Bhatti, Sheraz Ahmed

2018-03-01

A presence of mycotoxins in feed is one of the most alarming issues in the poultry feed industry. Ochratoxins, produced by several Aspergillus and Penicillium species, are important mycotoxin regarding the health status of poultry birds. Ochratoxins are further classified into to several subtypes (A, B, C, etc) depending on their chemical structures, but ochratoxin A (OTA) is considered the most important and toxic. Bentonite clay, belonging to phyllosilicates and formed from weathering of volcanic ashes, has adsorbent ability for several mycotoxins. The present study was designed to study the effects of bentonite clay upon OTA-induced immunosuppression in broiler chicks. For this, 480 day-old broiler chicks were procured from a local hatchery and then different combinations of OTA (0.15, 0.3, or 1.0 mg/kg) and bentonite clay (5, 10, and 20 g/kg) were incorporated into their feed. At 13, 30, and 42 days of age, parameters such as antibody responses to sheep red blood cells, in situ lymphoproliferative responses to mitogen (PHA-P), and in situ phagocytic activity (i.e., via carbon clearance) were determined respectively. The results indicated there was a significant reduction of total antibody and immunoglobulin titres, lymphoproliferative responses, and phagocytic potential in OTA-treated birds, suggesting clear immunosuppression by OTA in birds in a dose-dependent manner. These results were also significantly lower in all combination groups (OTA with bentonite clay), suggesting few to no effects of feeding bentonite clay upon OTA- induced alterations in different immune parameters.

Multi-walled carbon nanotubes as solid-phase extraction sorbents for simultaneous determination of type A trichothecenes in maize, wheat and rice by ultra-high performance liquid chromatography-tandem mass spectrometry.

PubMed

Dong, Maofeng; Si, Wenshuai; Jiang, Keqiu; Nie, Dongxia; Wu, Yongjiang; Zhao, Zhihui; De Saeger, Sarah; Han, Zheng

2015-12-04

A solid-phase extraction (SPE) procedure using multi-walled carbon nanotubes (MWCNTs) as sorbents coupled with ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) was developed for simultaneous determination of four type A trichothecenes in maize, wheat and rice for the first time. Several key parameters including the composition of sample loading solutions, washing and elution solvents were thoroughly investigated to achieve optimal SPE recoveries and efficiency. Performance of the MWCNTs materials was significantly affected by pH, and after optimization, n-hexane and 5% methanol aqueous solution as the washing solutions and methanol containing 1% formic acid as the elution solvent presented an excellent purification efficiency for the four targets in the different matrices. The method was validated by determining the linearity (R(2)≥0.992), recovery (73.4-113.7%), precision (1.2-17.1%) and sensitivity (limit of quantification in the range of 0.02-0.10μg/kg), and was further applied for simultaneous determination of type A trichothecenes in 30 samples. Although low contamination levels of type A trichothecenes in wheat, maize and rice were observed revealing mitigated risks to consumers in Shanghai, China, the developed method has proven to be a valuable tool for type A trichothecenes monitoring in complex crop matrices. Copyright © 2015 Elsevier B.V. All rights reserved.

Use of Nucleic Acid-Protein Cytophotometry and Cytointerferometry in Characterizing Cellular Mode of Trichothecene Toxicant Action

DTIC Science & Technology

1986-10-01

dose of 0.5 mg/k, the predominant bistopathic alterations included serosal petechial hemorrhages, moderate edema of the submucosa with Larea~e in...by a disruption of endothelial cell membranes, pycnotic nuclei, presence of edema in the interstitium and petechial hemorrhages within 2 h after T-2...Townsend. 1982. Inhibition, by trichothecene antibiotics, of brain protein synthesis and fever in rabbits. J. Physiol., Lond. 322:447. 21. Yarom, R

Effect of pH and solvent on the fluorescence spectroscopy of ochratoxin A

USDA-ARS?s Scientific Manuscript database

Ochratoxin A is a potential contaminant of agricultural commodities that is produced by Aspergillus and Penicillium species. Although this toxin has been observed at very low levels in commodities, it can reach levels of concern under certain conditions. It exhibits deleterious effects on animals an...

Presence of Multiple Mycotoxins and Other Fungal Metabolites in Native Grasses from a Wetland Ecosystem in Argentina Intended for Grazing Cattle

PubMed Central

Nichea, María J.; Palacios, Sofia A.; Chiacchiera, Stella M.; Sulyok, Michael; Krska, Rudolf; Chulze, Sofia N.; Torres, Adriana M.; Ramirez, María L.

2015-01-01

The aim of this study was to evaluate the occurrence of several fungal metabolites, including mycotoxins in natural grasses (Poaceae) intended for grazing cattle. A total number of 72 and 77 different metabolites were detected on 106 and 69 grass samples collected during 2011 and 2014, respectively. A total of 60 metabolites were found across both years. Among the few mycotoxins considered toxic for ruminants, no samples of natural grasses were contaminated with aflatoxins, ochratoxin A, ergot alkaloids, and gliotoxin, among others. However, we were able to detect important metabolites (toxic to ruminants) such as type A trichothecenes, mainly T-2 toxin and HT-2 toxin (up to 5000 µg/kg each), and zearalenone (up to 2000 µg/kg), all at very high frequencies and levels. Other fungal metabolites that were found to be prevalent were other Fusarium metabolites like beauvericin, equisetin and aurofusarin, metabolites produced by Alternaria spp., sterigmatocystin and its precursors and anthrachinone derivatives. It is important to point out that the profile of common metabolites was shared during both years of sampling, and also that the occurrence of important metabolites is not a sporadic event. Considering that this area of temperate grassland is used for grazing cattle all year long due to the richness in palatable grasses (Poaceae), the present work represents a starting point for further studies on the occurrence of multi-mycotoxins in natural grasses in order to have a complete picture of the extent of cattle exposure. Also, the present study shows that the presence of zeranol in urine of beef cattle may not be a consequence of illegal use of this banned substance, but the product of the natural occurrence of zearalenone and α-zearalenol in natural grasses intended for cattle feeding. PMID:26308052

Mass Spectral Investigations on Toxins. 2. Simultaneous Detection and Quantification of Ultra-Trace Levels of Simple Trichothecenes in Environmental and Fermentation Samples by Gas Chromatographic/Negative Ion Chemical Ionization-Mass Spectrometric Techniques

DTIC Science & Technology

1987-01-01

due to interferences in the pollen. However, the identity of the interferents is presently unknown. A dried papaya leaf was treated with 10 ml of warm...Known amounts of DON, DAS, and T-2 were spiked on a blank (trichothecene-free) papaya leaf and left exposed in a bottle for 1 year. At the end of the year...Simple Trichothecenes from Leaf Sample after Prolonged Exposure ............... 35 12 Sample Analysis .............................. .... 37 6 MASS

Modulation of Ergot Alkaloids in a Grass-Endophyte Symbiosis by Alteration of mRNA Concentrations of an Ergot Alkaloid Synthesis Gene.

PubMed

Mulinti, Prashanthi; Florea, Simona; Schardl, Christopher L; Panaccione, Daniel G

2016-06-22

The profile of ergot alkaloids in perennial ryegrass (Lolium perenne) containing the endophytic fungus Epichloë typhina × festucae includes high concentrations of the early pathway metabolites ergotryptamine and chanoclavine-I in addition to the pathway end-product ergovaline. Because these alkaloids differ in activity, we investigated strategies to alter their relative concentrations. An RNAi-based approach reduced the concentration of mRNA from the gene easA, which encodes an enzyme required for a ring closure that separates ergotryptamine and chanoclavine-I from ergovaline. Lower easA mRNA concentrations correlated with lower concentrations of ergovaline and higher concentrations of ergotryptamine and chanoclavine-I. Overexpression of easA led to higher concentrations of ergovaline in leaf blades but not in pseudostems; concentrations of the early pathway metabolites were not altered in overexpression strains. The data indicate that altering the concentration of mRNA from a single gene can change alkaloid flux, but the magnitude of the change was limited and variable.

Deoxynivalenol, deoxynivalenol-3-glucoside, and enniatins: the major mycotoxins found in cereal-based products on the Czech market.

PubMed

Malachova, Alexandra; Dzuman, Zbynek; Veprikova, Zdenka; Vaclavikova, Marta; Zachariasova, Milena; Hajslova, Jana

2011-12-28

Fusarium toxins, Alternaria toxins, and ergot alkaloids represent common groups of mycotoxins that can be found in cereals grown under temperate climatic conditions. Because most of them are chemically and thermally stable, these toxic fungal secondary metabolites might be transferred from grains into the final products. To get information on the commensurate contamination of various cereal-based products collected from the Czech retail market in 2010, the occurrence of "traditional" mycotoxins such as groups of A and B trichothecenes and zearalenone, less routinely determined Alternaria toxins (alternariol, alternariol monomethyl ether and altenuene), ergot alkaloids (ergosine, ergocryptine, ergocristine, and ergocornine) and "emerging" mycotoxins (enniatins A, A1, B, and B1 and beauvericin) were monitored. In a total 116 samples derived from white flour and mixed flour, breakfast cereals, snacks, and flour, only trichothecenes A and B and enniatins were found. Deoxynivalenol was detected in 75% of samples with concentrations ranging from 13 to 594 μg/kg, but its masked form, deoxynivalenol-3-β-d-glucoside, has an even higher incidence of 80% of samples, and concentrations ranging between 5 and 72 μg/kg were detected. Nivalenol was found only in three samples at levels of 30 μg/kg. For enniatins, all of the samples investigated were contaminated with at least one of four target enniatins. Enniatin A was detected in 97% of samples (concentration range of 20-2532 μg/kg) followed by enniatin B with an incidence in 91% of the samples (concentration range of 13-941 μg/kg) and enniatin B1 with an incidence of 80% in the samples tested (concentration range of 8-785 μg/kg). Enniatin A1 was found only in 44% of samples at levels ranging between 8 and 851 μg/kg.

Effect of trichothecene production on the plant defense response and fungal physiology: overexpression of Trichoderma arundinaceum tri4 gene in T. harzianum

USDA-ARS?s Scientific Manuscript database

Trichothecenes are fungal sesquiterpenoid compounds, the majority of which have phytotoxic activity. They contaminate food and feed stocks, resulting in potential harm to animals and human beings. Trichoderma brevicompactum and T. arundinaceum produce trichodermin and harzianum A (HA), respectively,...

Effects of Ochratoxin A on Livestock Production

PubMed Central

Battacone, Gianni; Nudda, Anna; Pulina, Giuseppe

2010-01-01

Ochratoxin A (OTA) contamination often causes large economic losses on livestock production. The intake of feed contaminated by OTA also represents a potential risk for animal health and a food safety issue due to the transfer of the toxin through the food chain to humans. The aim of this paper is to review the available literature on: (1) the frequency and degree of occurrence of OTA in different feedstuffs; (2) the toxicological effects of OTA intake on the performance of the main livestock (i.e., poultry, swine, cattle, goats and sheep); and (3) the transfer of OTA, or its metabolites, from animal feed into animal products such as milk, meat and eggs. PMID:22069661

Polyaniline Langmuir-Blodgett film based aptasensor for ochratoxin A detection.

PubMed

Prabhakar, Nirmal; Matharu, Zimple; Malhotra, B D

2011-06-15

Ochratoxin A (OTA) produced by Aspergillus Ochraceus and Penicillium verrucosum is a very dangerous toxin due to its toxic effects in human beings and its presence in a wide range of food products and cereals. A Langmuir-Blodgett (polyaniline (PANI)-stearic acid (SA)) film based highly sensitive and robust impedimetric aptasensor has been developed for ochratoxin A (OTA) detection. DNA Aptamer (Apt-DNA) specific to OTA has been covalently immobilized onto mixed Langmuir-Blodgett (LB) monolayer comprising of PANI-SA deposited onto indium tin-oxide (ITO) coated glass plates. This Apt-DNA/PANI-SA/ITO aptaelectrode has been characterized using scanning electron microscopy, Fourier transform-infrared spectroscopy, contact angle measurements, cyclic voltammetry and electrochemical impedance spectroscopy, respectively. The Apt-DNA/PANI-SA/ITO aptasensor shows detection of OTA by electrochemical impedance spectroscopy in the linear range of 0.0001 μg/ml (0.1 ng/ml) to 0.01 μg/ml (10 ng/ml) and 1 μg/ml-25 μg/ml with detection limit of 0.1 ng/ml in 15 min. The Apt-DNA/PANI-SA/ITO aptasensor can be reused ∼13 times. The binding or affinity constant (K(a)) of aptamer with OTA, calculated using Langmuir adsorption isotherm, is found be 1.21×10(7) M(-1). Copyright © 2011 Elsevier B.V. All rights reserved.

Differences in the regulation of ochratoxin A by the HOG pathway in Penicillium and Aspergillus in response to high osmolar environments.

PubMed

Stoll, Dominic; Schmidt-Heydt, Markus; Geisen, Rolf

2013-07-19

Penicillium verrucosum, P. nordicum and Aspergillus carbonarius are three important ochratoxin A producing species. P. verrucosum is in addition able to produce citrinin. It has been shown earlier that P. nordicum is adapted to NaCl rich environments like salt rich dry cured foods or even salines. In this organism, the biosynthesis of ochratoxin A plays an adaptive role in this habitat. P. verrucosum generally can be found on cereals, but occasionally also on salt rich dry cured foods. In contrast A. carbonarius usually cannot be found in NaCl rich environments, but it occurs in another environment with high concentration of solutes, e.g., in sugar rich substrates like grapes and grape juices. Usually osmotic challenging conditions activate the HOG MAP kinase signal cascade, which in turn activates various osmo-regulated genes. In the current analysis, it could be demonstrated that in case of P. nordicum and P. verrucosum the NaCl induced production of ochratoxin A is correlated to the phosphorylation status of the HOG MAP kinase. Just the opposite was true for A. carbonarius. In this case, also higher amounts of NaCl in the medium lead to an increased phosphorylation status of HOG, but no increase in ochratoxin biosynthesis was observed. In contrast to the Penicillia, higher NaCl concentrations lead to a rapid cessation of growth by A. carbonarius. High glucose concentrations have much less impact on growth and the phosphorylation of HOG.

Molecular characterization of black Aspergillus species from onion and their potential for ochratoxin A and fumonisin B2 production.

PubMed

Gherbawy, Youssuf; Elhariry, Hesham; Kocsubé, Sándor; Bahobial, Abdulaziz; Deeb, Bahig El; Altalhi, Abdulla; Varga, János; Vágvölgyi, Csaba

2015-05-01

Onion bulbs can become contaminated with various molds during the storage period, the most important causal agents being black aspergilli (Aspergillus section Nigri). Taxonomic studies have revealed that this group of Aspergillus contains many species that cannot be reliably identified using standard morphological methods. Therefore, it is necessary to define the fungus causing this problem in the onion exactly, especially since some species assigned to section Nigri are well known as ochratoxin and/or fumonisin producers. Sixty fungal isolates belonging to 10 fungal genera were isolated from 40 onion samples originated from the Taif region in Saudi Arabia. Black aspergilli were detected in 37 onion samples. Using primer pairs (awaspec and Cmd6) designed based on partial calmodulin gene sequence data, 37 isolates were identified as A. welwitschiae. The ochratoxin A and fumonisin B2 contents of the onion samples were examined. No ochratoxins were detected in the collected samples, while fumonisin B2 was detected in 37.5% of the onion samples. Eighteen of 37 isolates of Aspergillus welwitschiae were recognized as potential producers for fumonisin B2. Multiplex polymerase chain reactions designed to detect biosynthetic genes of fumonisins confirmed these results.

Occurrence of ochratoxin A in cocoa products and chocolate.

PubMed

Serra Bonvehí, Josep

2004-10-06

In this work, the occurrence of ochratoxin A (OTA) in 170 samples of cocoa products of different geographical origins was studied. An immunoaffinity column with HPLC separation was developed to quantify low levels of OTA in cocoa bean, cocoa cake, cocoa mass, cocoa nib, cocoa powder, cocoa shell, cocoa butter, chocolate, and chocolate cream with >80% recoveries. The method was validated by performing replicate analyses of uncontaminated cocoa material spiked at three different levels of OTA (1, 2, and 5 microg/kg). The data obtained were related on the acceptable safe daily exposure for OTA. The highest levels of OTA were detected in roasted cocoa shell and cocoa cake (0.1-23.1 microg/kg) and only at minor levels in the other cocoa products. Twenty-six cocoa and chocolate samples were free from detectable OTA (<0.10 microg/kg). In roasted cocoa powder 38.7% of the samples analyzed contained OTA at levels ranging from 0.1 to 2 microg/kg, and 54.8% was contaminated at >2 microg/kg (and 12 samples at >3 microg/kg). Ochratoxin A was detected in cocoa bean at levels from 0.1 to 3.5 microg/kg, the mean concentration being 0.45 microg/kg; only one sample exceeded 2 microg/kg (4.7%). In contrast, 51.2% of cocoa cake samples contained OTA at levels > or =2 microg/kg, among which 16 exceeded 5 microg/kg (range of 5-9 microg/kg). These results indicate that roasted cocoa powder is not a major source of OTA in the diet.

Aflatoxin and ochratoxin A content of spices in Hungary.

PubMed

Fazekas, B; Tar, A; Kovács, M

2005-09-01

In October and November 2004, 91 spice samples (70 ground red pepper, six black pepper, five white pepper, five spice mix and five chilli samples), the majority of which originated from commercial outlets, were analysed for aflatoxins B1, B2, G1 and G2 (AFB1, AFB2, AFG1, AFG2) and ochratoxin A (OTA) content by high-performance liquid chromatography (HPLC) after immunoaffinity column clean-up. Eighteen of the 70 ground red pepper samples contained AFB1, seven of them in a concentration exceeding the 'maximum level' of 5 microg kg(-1) (range 6.1-15.7 microg kg(-1)). Of the other spices assayed, the AFB1 contamination of one chilli sample exceeded 5 microg kg(-1) (8.1 microg kg(-1)). Thirty-two of the 70 ground red pepper samples contained OTA, eight of them in a concentration exceeding the 10 microg kg(-1) 'maximum level' (range 10.6-66.2 microg kg(-1)). One chilli sample was contaminated with OTA at 2.1 microg kg(-1). The AFB1 and OTA contamination of ground red pepper exceeding the 'maximum level' (5 and 10 microg kg(-1), respectively) was obviously the consequence of mixing imported ground red pepper batches heavily contaminated with AFB1 and OTA with red pepper produced in Hungary. This case calls attention to the importance of consistently screening imported batches of ground red pepper for aflatoxin and ochratoxin A content and strictly prohibiting the use of batches containing mycotoxin concentrations exceeding the maximum permitted level.

Estimated dietary exposure to principal food mycotoxins from the first French Total Diet Study.

PubMed

Leblanc, J-C; Tard, A; Volatier, J-L; Verger, P

2005-07-01

This study reports estimates on dietary exposure from the first French Total Diet Study (FTDS) and compares these estimates with both existing tolerable daily intakes for these toxins and the intakes calculated during previous French studies. To estimate the dietary exposure of the French population to the principal mycotoxins in the French diet (as consumed), 456 composite samples were prepared from 2280 individual samples and analysed for aflatoxins, ochratoxin A, trichothecenes, zearalenone, fumonisins and patulin. Average and high percentile intakes were calculated taking account of different eating patterns for adults, children and vegetarians. The results showed that contaminant levels observed in the foods examined 'as consumed' complied fully with current European legislation. However, particular attention needs to be paid to the exposure of specific population groups, such as children and vegans/macrobiotics, who could be exposed to certain mycotoxins in quantities that exceed the tolerable or weekly daily intake levels. This observation is particularly relevant with respect to ochratoxin A, deoxynivalenol and zearalenone. For these mycotoxins, cereals and cereal products were the main contributors to high exposure.

Proteomic analysis of a mutant of Trichoderma arundinaceum impaired in the trichothecene biosynthesis reveals a systemic function of these compounds in the fungal physiology

USDA-ARS?s Scientific Manuscript database

Trichothecenes are sesquiterpene mycotoxins produced by several fungal genera including Fusarium, Trichothecium, Myrothecium, Stachybotrys, and Trichoderma. These toxins have attracted great attention because they are frequent contaminants of food and animal feed, and can be easily absorbed by anim...

Neurotoxic, inflammatory, and mucosecretory responses in the nasal airways of mice repeatedly exposed to the macrocyclic trichothecene mycotoxin roridin A: dose-response and persistence of injury.

PubMed

Corps, Kara N; Islam, Zahidul; Pestka, James J; Harkema, Jack R

2010-04-01

Macrocyclic trichothecene mycotoxins encountered in water-damaged buildings have been suggested to contribute to illnesses of the upper respiratory tract. Here, the authors characterized the adverse effects of repeated exposures to roridin A (RA), a representative macrocyclic trichothecene, on the nasal airways of mice and assessed the persistence of these effects. Young, adult, female C57BL/6 mice were exposed to single daily, intranasal, instillations of RA (0.4, 2, 10, or 50 microg/kg body weight [bw]) in saline (50 microl) or saline alone (controls) over 3 weeks or 250 microg/kg RA over 2 weeks. Histopathologic, immunohistochemical, and morphometric analyses of nasal airways conducted 24 hr after the last instillation revealed that the lowest-effect level was 10 microg/kg bw. RA exposure induced a dose-dependent, neutrophilic rhinitis with mucus hypersecretion, atrophy and exfoliation of nasal transitional and respiratory epithelium, olfactory epithelial atrophy and loss of olfactory sensory neurons (OSNs). In a second study, the persistence of lesions in mice instilled with 250 microg/kg bw RA was assessed. Nasal inflammation and excess luminal mucus were resolved after 3 weeks, but OSN loss was still evident in olfactory epithelium (OE). These results suggest that nasal inflammation, mucus hypersecretion, and olfactory neurotoxicity could be important adverse health effects associated with short-term, repeated, airborne exposures to macrocyclic trichothecenes.

Acute exposure to ergot alkaloids from endophyte-infected tall fescue does not alter absorptive or barrier function of the isolated bovine ruminal epithelium.

PubMed

Foote, A P; Penner, G B; Walpole, M E; Klotz, J L; Brown, K R; Bush, L P; Harmon, D L

2014-07-01

Ergot alkaloids in endophyte-infected (Neotyphodium coenophialum) tall fescue (Lolium arundinaceum) have been shown to cause a reduction in blood flow to the rumen epithelium as well as a decrease in volatile fatty acids (VFA) absorption from the washed rumen of steers. Previous data also indicates that incubating an extract of endophyte-infected tall fescue seed causes an increase in the amount of VFA absorbed per unit of blood flow, which could result from an alteration in the absorptive or barrier function of the rumen epithelium. An experiment was conducted to determine the acute effects of an endophyte-infected tall fescue seed extract (EXT) on total, passive or facilitated acetate and butyrate flux across the isolated bovine rumen as well as the barrier function measured by inulin flux and tissue conductance (G t ). Flux of ergovaline across the rumen epithelium was also evaluated. Rumen tissue from the caudal dorsal sac of Holstein steers (n=6), fed a common diet, was collected and isolated shortly after slaughter and mounted between two halves of Ussing chambers. In vitro treatments included vehicle control (80% methanol, 0.5% of total volume), Low EXT (50 ng ergovaline/ml) and High EXT (250 ng ergovaline/ml). Results indicate that there is no effect of acute exposure to ergot alkaloids on total, passive or facilitated flux of acetate or butyrate across the isolate bovine rumen epithelium (P>0.51). Inulin flux (P=0.16) and G t (P>0.17) were not affected by EXT treatment, indicating no alteration in barrier function due to acute ergot alkaloid exposure. Ergovaline was detected in the serosal buffer of the High EXT treatment indicating that the flux rate is ~0.25 to 0.44 ng/cm2 per hour. Data indicate that specific pathways for VFA absorption and barrier function of the rumen epithelium are not affected by acute exposure to ergot alkaloids from tall fescue at the concentrations tested. Ergovaline has the potential to be absorbed from the rumen of cattle that

The induction of mycotoxins by trichothecene producing Fusarium species.

PubMed

Lowe, Rohan; Jubault, Mélanie; Canning, Gail; Urban, Martin; Hammond-Kosack, Kim E

2012-01-01

In recent years, many Fusarium species have emerged which now threaten the productivity and safety of small grain cereal crops worldwide. During floral infection and post-harvest on stored grains the Fusarium hyphae produce various types of harmful mycotoxins which subsequently contaminate food and feed products. This article focuses specifically on the induction and production of the type B sesquiterpenoid trichothecene mycotoxins. Methods are described which permit in liquid culture the small or large scale production and detection of deoxynivalenol (DON) and its various acetylated derivatives. A wheat (Triticum aestivum L.) ear inoculation assay is also explained which allows the direct comparison of mycotoxin production by species, chemotypes and strains with different growth rates and/or disease-causing abilities. Each of these methods is robust and can be used for either detailed time-course studies or end-point analyses. Various analytical methods are available to quantify the levels of DON, 3A-DON and 15A-DON. Some criteria to be considered when making selections between the different analytical methods available are briefly discussed.

Acute exposure to ergot alkaloids from endophyte-infected tall fescue does not alter absorptive or barrier function of the isolated ruminal epithelium

USDA-ARS?s Scientific Manuscript database

Ergot alkaloids in endophyte-infected (Neotyphodium coenophialum) tall fescue (Lolium arundinaceum) have been shown to cause a reduction in blood flow to the rumen epithelium as well as a decrease in VFA absorption from the washed rumen of steers. Previous data also indicates that incubating an extr...

Cetylpyridinium chloride functionalized silica-coated magnetite microspheres for the solid-phase extraction and pre-concentration of ochratoxin A from environmental water samples with high-performance liquid chromatographic analysis.

PubMed

Hu, Meihua; Huang, Pengcheng; Suo, Lili; Wu, Fangying

2017-06-01

A new method based on cetylpyridinium chloride coated ferroferric oxide/silica magnetic microspheres as an efficient solid-phase adsorbent was developed for the extraction and enrichment of ochratoxin A. The determination of ochratoxin A was obtained by high-performance liquid chromatography with fluorescence detection. In the presence of cetylpyridinium chloride, the adsorption capacity of ferroferric oxide/silica microspheres was 5.95 mg/g for ochratoxin A. The experimental parameters were optimized, including the amounts of ferroferric oxide/silica microspheres (20 mg) and cetylpyridinium chloride (0.18 mL, 0.5 mg/mL), pH value of media (9), ultrasonic time (5 min), elution solvent and volume [2(1 + 1) mL (washed twice, 1 mL each time) 1% acetic acid acetonitrile]. Under optimal experiment conditions, ochratoxin A had good linearity in the range of 2.5-250.0 ng/L in water samples with correlation coefficient of the calibration curve 0.9995. The limit of detection for ochratoxin A was 0.83 ng/L, and the recoveries were 89.8-96.8% with the relative standard deviation of 1.5-3.5% in environmental water samples. Furthermore, ferroferric oxide/silica microspheres show excellent reusability during extraction procedures for no less than six times. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Ochratoxin a and mitotic disruption: mode of action analysis of renal tumor formation by ochratoxin A.

PubMed

Mally, Angela

2012-06-01

The mycotoxin and food contaminant ochratoxin A (OTA) is a potent renal carcinogen in rodents, but its mode of action (MoA) is still poorly defined. In 2006, the European Food Safety Authority concluded that there is a "lack of evidence for the existence of OTA-DNA adducts" and thus insufficient evidence to establish DNA reactivity as a MoA for tumor formation by OTA. In reviewing the available database on OTA toxicity, a MoA for renal carcinogenicity of OTA is developed that involves a combination of genetic instability and increased proliferative drive as consequences of OTA-mediated disruption of mitosis, whereby the organ- and site-specificity of tumor formation by OTA is determined by selective renal uptake of OTA into the proximal tubule epithelium. The proposed MoA is critically assessed with respect to concordance of dose-response of the suggested key events and tumor formation, their temporal association, consistency, and biological plausibility. Uncertainties, data gaps and needs for further research are highlighted.

Mycotoxins in foods in Lower Saxony (Germany): results of official control analyses performed in 2009.

PubMed

Reinhold, Lilli; Reinhardt, Katja

2011-05-01

In this presentation, the mycotoxin levels-as analysed by the analytical centre for mycotoxin surveillance of the state food laboratory (LAVES Braunschweig)-for approximately 500 food samples are reported. The samples were collected in the year 2009 at retail in the German federal state of Lower Saxony. Aflatoxin and ochratoxin A were analysed in dried fruits, spices, cereals and tree nuts. Ochratoxin A was detected in all samples of dried vine fruits, at levels up to 8.1 μg/kg. Aflatoxins and ochratoxin A were also found in nutmeg and curry powder: the maximum regulatory levels for aflatoxins were exceeded in 25% of the nutmeg samples. Nearly all samples of basmati rice contained aflatoxins, although at levels below the maximum regulatory level in all but one sample. Aflatoxins were also detected in about 50% of hazelnut samples, in 20% of the samples the maximum levels was exceeded (maximum 23.2 μg/kg). In contrast, aflatoxin contents in pistachios were surprisingly low. Fusarium toxins were analysed in cereals and cereal products such as flour, bread, and pasta. Deoxynivalenol (DON) was the predominant toxin found in these samples: DON was found in about 40% of the samples, although the maximum levels were not exceeded (max. 418 μg/kg). Fumonisins (FBs) and zearalenone (ZEA) were specifically analysed in maize products (snacks, flour and oil). Most of these samples (80%) were positive, but at levels not exceeding the maximum levels. Maximum levels were 98 μg/kg (ZEA) and 577 μg/kg (sum of FB1 and FB2). Ergot alkaloids (six major alkaloids) were analysed in rye flour, and approximately 50% were positive. The highest concentration of ergot alkaloids was 1,063 μg/kg; the predominant alkaloids were ergotamine and ergocristine. In conclusion, the results indicate that continuous and efficient control measures for mycotoxins in a wide range of critical foods are necessary to ensure compliance with maximum levels. Although the mycotoxin levels in the vast

Aerobic De-Epoxydation of Trichothecene Mycotoxins by a Soil Bacterial Consortium Isolated Using In Situ Soil Enrichment.

PubMed

He, Wei-Jie; Yuan, Qing-Song; Zhang, You-Bing; Guo, Mao-Wei; Gong, An-Dong; Zhang, Jing-Bo; Wu, Ai-Bo; Huang, Tao; Qu, Bo; Li, He-Ping; Liao, Yu-Cai

2016-09-24

Globally, the trichothecene mycotoxins deoxynivalenol (DON) and nivalenol (NIV) are among the most widely distributed mycotoxins that contaminate small grain cereals. In this study, a bacterial consortium, PGC-3, with de-epoxydation activity was isolated from soil by an in situ soil enrichment method. Screening of 14 soil samples that were sprayed with DON revealed that 4 samples were able to biotransform DON into de-epoxydized DON (dE-DON). Among these, the PGC-3 consortium showed the highest and most stable activity to biotransform DON into dE-DON and NIV into dE-NIV. PGC-3 exhibited de-epoxydation activity at a wide range of pH (5-10) and temperatures (20-37 °C) values under aerobic conditions. Sequential subculturing with a continued exposure to DON substantially reduced the microbial population diversity of this consortium. Analyses of the 16S rDNA sequences indicated that PGC-3 comprised 10 bacterial genera. Among these, one species, Desulfitobacterium, showed a steady increase in relative abundance, from 0.03% to 1.55% (a 52-fold increase), as higher concentrations of DON were used in the subculture media, from 0 to 500 μg/mL. This study establishes the foundation to further develop bioactive agents that can detoxify trichothecene mycotoxins in cereals and enables for the characterization of detoxifying genes and their regulation.

Aerobic De-Epoxydation of Trichothecene Mycotoxins by a Soil Bacterial Consortium Isolated Using In Situ Soil Enrichment

PubMed Central

He, Wei-Jie; Yuan, Qing-Song; Zhang, You-Bing; Guo, Mao-Wei; Gong, An-Dong; Zhang, Jing-Bo; Wu, Ai-Bo; Huang, Tao; Qu, Bo; Li, He-Ping; Liao, Yu-Cai

2016-01-01

Globally, the trichothecene mycotoxins deoxynivalenol (DON) and nivalenol (NIV) are among the most widely distributed mycotoxins that contaminate small grain cereals. In this study, a bacterial consortium, PGC-3, with de-epoxydation activity was isolated from soil by an in situ soil enrichment method. Screening of 14 soil samples that were sprayed with DON revealed that 4 samples were able to biotransform DON into de-epoxydized DON (dE-DON). Among these, the PGC-3 consortium showed the highest and most stable activity to biotransform DON into dE-DON and NIV into dE-NIV. PGC-3 exhibited de-epoxydation activity at a wide range of pH (5–10) and temperatures (20–37 °C) values under aerobic conditions. Sequential subculturing with a continued exposure to DON substantially reduced the microbial population diversity of this consortium. Analyses of the 16S rDNA sequences indicated that PGC-3 comprised 10 bacterial genera. Among these, one species, Desulfitobacterium, showed a steady increase in relative abundance, from 0.03% to 1.55% (a 52-fold increase), as higher concentrations of DON were used in the subculture media, from 0 to 500 μg/mL. This study establishes the foundation to further develop bioactive agents that can detoxify trichothecene mycotoxins in cereals and enables for the characterization of detoxifying genes and their regulation. PMID:27669304

A Review of the Diagnosis and Treatment of Ochratoxin A Inhalational Exposure Associated with Human Illness and Kidney Disease including Focal Segmental Glomerulosclerosis

PubMed Central

Hope, Janette H.; Hope, Bradley E.

2012-01-01

Ochratoxin A (OTA) exposure via ingestion and inhalation has been described in the literature to cause kidney disease in both animals and humans. This paper reviews Ochratoxin A and its relationship to human health and kidney disease with a focus on a possible association with focal segmental glomerulosclerosis (FSGS) in humans. Prevention and treatment strategies for OTA-induced illness are also discussed, including cholestyramine, a bile-acid-binding resin used as a sequestrant to reduce the enterohepatic recirculation of OTA. PMID:22253638

Impedimetric Aptasensor for Ochratoxin A Determination Based on Au Nanoparticles Stabilized with Hyper-Branched Polymer

PubMed Central

Evtugyn, Gennady; Porfireva, Anna; Stepanova, Veronika; Kutyreva, Marianna; Gataulina, Alfiya; Ulakhovich, Nikolay; Evtugyn, Vladimir; Hianik, Tibor

2013-01-01

An impedimetric aptasensor for ochratoxin A (OTA) detection has been developed on the base of a gold electrode covered with a new modifier consisting of electropolymerized Neutral Red and a mixture of Au nanoparticles suspended in the dendrimeric polymer Botlorn H30®. Thiolated aptamer specific to OTA was covalently attached to Au nanoparticles via Au-S bonding. The interaction of the aptamer with OTA induced the conformational switch of the aptamer from linear to guanine quadruplex form followed by consolidation of the surface layer and an increase of the charge transfer resistance. The aptasensor makes it possible to detect from 0.1 to 100 nM of OTA (limit of detection: 0.02 nM) in the presence of at least 50 fold excess of ochratoxin B. The applicability of the aptasensor for real sample assay was confirmed by testing spiked beer samples. The recovery of 2 nM OTA was found to be 70% for light beer and 78% for dark beer. PMID:24287535

Structural and Functional Characterization of the TRI101 Trichothecene 3-O-Acetyltransferase from Fusarium sporotrichioides and Fusarium graminearum: KINETIC INSIGHTS TO COMBATING FUSARIUM HEAD BLIGHT

DOE Office of Scientific and Technical Information (OSTI.GOV)

Garvey, Graeme S.; McCormick, Susan P.; Rayment, Ivan

2008-06-30

Fusarium head blight (FHB) is a plant disease with serious economic and health impacts. It is caused by fungal species belonging to the genus Fusarium and the mycotoxins they produce. Although it has proved difficult to combat this disease, one strategy that has been examined is the introduction of an indigenous fungal protective gene into cereals such as wheat barley and rice. Thus far the gene of choice has been tri101 whose gene product catalyzes the transfer of an acetyl group from acetyl coenzyme A to the C3 hydroxyl moiety of several trichothecene mycotoxins. In vitro this has been shownmore » to reduce the toxicity of the toxins by {approx}100-fold but has demonstrated limited resistance to FHB in transgenic cereal. To understand the molecular basis for the differences between in vitro and in vivo resistance the three-dimensional structures and kinetic properties of two TRI101 orthologs isolated from Fusarium sporotrichioides and Fusarium graminearum have been determined. The kinetic results reveal important differences in activity of these enzymes toward B-type trichothecenes such as deoxynivalenol. These differences in activity can be explained in part by the three-dimensional structures for the ternary complexes for both of these enzymes with coenzyme A and trichothecene mycotoxins. The structural and kinetic results together emphasize that the choice of an enzymatic resistance gene in transgenic crop protection strategies must take into account the kinetic profile of the selected protein.« less

SOLVENT COMPARISON IN THE ISOLATION, SOLUBILIZATION, AND TOXICITY OF STACHYBOTRYS CHARTARUM SPORE TRICHOTHECENE MYCOTOXINS IN AN ESTABLISHED IN VITRO LUMINESCENCE PROTEIN TRANSLATION INHIBITION ASSAY

EPA Science Inventory

It is well known that non-viable mold contaminants such as macrocyclic trichothecene mycotoxins of Stachybotrys chartarum are highly toxinigenic to humans. However, there is no agreed upon method of recovering native mycotoxin. The purpose of this study was to provide quantitativ...

GLUTAMIC DECARBOXYLASE OF ERGOT, CLAVICEPS PURPUREA

PubMed Central

Anderson, John A.; Cheldelin, Vernon H.; King, Tsoo E.

1961-01-01

Anderson, John A. (Oregon State University, Corvallis), Vernon H. Cheldelin, and Tsoo E. King. Glutamic decarboxylase of ergot, Claviceps purpurea. J. Bacteriol. 82:354–358. 1961.—l-Glutamic acid is the only naturally occurring amino acid which can be decarboxylated by cell-free extracts of Claviceps purpurea. This decarboxylase was partially purified and the properties of the enzyme studied. The specific activity of the purified preparation was 111 μliters per 10 min per mg of protein. The products formed, stability, inhibition, stimulation of activity with pyridoxal phosphate, and pH activity curve were typical of l-glutamic decarboxylase in Escherichia coli and other microorganisms. The substrate constants at pH 4.6, 5.25, and 5.65 were 0.0169 m, 0.0174 m, and 0.0139 m, respectively. The respective maximal velocities at these pH values were 104, 104, and 90 μliters per 10 min. The pH optimum was 4.8 to 5.2. The enzyme was unstable below pH 4.5 and it was suggested that the fall in activity at the lower end of the pH curve was due to inactivation of the enzyme. The decrease in activity above pH 5.2 did not appear to be due to a change in affinity of enzyme for substrate but to a change of the enzyme-substrate complex into an inactive form. PMID:13683214

Studies on some feed additives and materials giving partial protection against the suppressive effect of ochratoxin A on egg production of laying hens.

PubMed

Stoev, Stoycho D

2010-06-01

The protective effects of various feed supplements against the harmful effect of ochratoxin A on egg production and sexual maturation of two-weeks old Plymouth Rock female chicks designed for laying hens were studied. A significant protective effect of the feed additives or materials: water extract of artichoke (WEA), sesame seed (SS), Roxazyme-G (RG) and l-beta phenylalanine (PHE) against the suppressive effect of ochratoxin A (OTA) on egg production of laying hens was found. A similar protection was also seen on the toxic effect of OTA on various internal organs of the same hens. A significant protection was found against the decrease of the weight or the quantity of eggs as well as against the delay of the beginning of the laying period of chicks, both of which were provoked by ochratoxin A. These protective effects were strongest in chicks treated with SS or WEA, but were slightest in chicks treated with l-beta PHE. Copyright 2009 Elsevier Ltd. All rights reserved.

A New Ochratoxin A Biodegradation Strategy Using Cupriavidus basilensis Őr16 Strain

PubMed Central

Krifaton, Csilla; Szoboszlay, Sándor; Kukolya, József; Szőke, Zsuzsanna; Kőszegi, Balázs; Albert, Mihály; Barna, Teréz; Mézes, Miklós; Kovács, Krisztina J.; Kriszt, Balázs

2014-01-01

Ochratoxin-A (OTA) is a mycotoxin with possibly carcinogenic and nephrotoxic effects in humans and animals. OTA is often found as a contaminant in agricultural commodities. The aim of the present work was to evaluate OTA-degrading and detoxifying potential of Cupriavidus basilensis ŐR16 strain. In vivo administration of OTA in CD1 male mice (1 or 10 mg/kg body weight for 72 hours or 0.5 mg/kg body weight for 21 days) resulted in significant elevation of OTA levels in the blood, histopathological alterations- and transcriptional changes in OTA-dependent genes (annexinA2, clusterin, sulphotransferase and gadd45 and gadd153) in the renal cortex. These OTA-induced changes were not seen in animals that have been treated with culture supernatants in which OTA was incubated with Cupriavidus basilensis ŐR16 strain for 5 days. HPLC and ELISA methods identified ochratoxin α as the major metabolite of OTA in Cupriavidus basilensis ŐR16 cultures, which is not toxic in vivo. This study has demonstrated that Cupriavidus basilensis ŐR16 efficiently degrade OTA without producing toxic adventitious metabolites. PMID:25302950

Development and validation of LC-MS/MS method for the determination of Ochratoxin A and its metabolite Ochratoxin α in poultry tissues and eggs.

PubMed

Paoloni, Angela; Solfrizzo, Michele; Bibi, Rita; Pecorelli, Ivan

2018-05-04

The objective of this study was to develop a liquid chromatography tandem mass spectrometry (LC-MS/MS) method for the determination of Ochratoxin A (OTA) and Ochratoxin α (OTα) in poultry tissues and eggs. The two toxins were extracted by a mixture of acetonitrile/water, purified with a reversed phase C18 solid phase extraction column (SPE) and determined by LC-MS/MS. The LC-MS/MS method performances were evaluated in terms of linearity in solvent and in matrix (ranged from 0.5 to 15.10 µg L -1 for OTA and from 0.60 to 17.85 µg L -1 for OTα), limit of detection (LOD), limit of quantitation (LOQ), specificity, accuracy and precision in repeatability conditions. Recovery experiments were performed by spiking poultry liver, kidney, muscle and eggs around 1 µg kg -1 and 10 µg kg -1 . LODs were 0.27 and 0.26 µg kg -1 while LOQs were fixed at 1.0 and 1.2 µg kg -1 for OTA and OTα, respectively. Main recoveries for OTA ranged from 82 to 109% and for OTα ranged from 55 to 89%. The values of within-laboratory relative standard deviation (RSD r ) were equal to or below 20%. Considering the results obtained and that all analytical performance criteria were fulfilled, the new extraction and purification method developed for OTA and OTα determination in animal tissues and eggs was found appropriate for control laboratories and research activities designed to ensure food safety.

Case Study: Recovery from ergot alkaloid-induced vasoconstriction for steers conditioned to grazing seedhead suppressed and unsuppressed pastures of toxic endophyte-infected tall fescue

USDA-ARS?s Scientific Manuscript database

Chemical seedhead suppression of toxic endophyte-infected (E+) tall fescue can enhance steer performance and mitigate the adverse effects of ergot alkaloids on cattle physiology; however, it is not known if seedhead suppression can mitigate alkaloid-induced vasoconstriction and improve post-graze pe...

Controlled study on the effect of pentoxifylline and an ergot alkaloid derivative on regional cerebral blood flow in patients with chronic cerebrovascular disease

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hartmann, A.; Tsuda, Y.

Regional cerebral blood flow (rCBF) in 90 patients with CBF decreased due to vascular diseases was studied by using the xenon 133 inhalation technique and a 32-detector setup. Whereas 30 patients received their standard basic therapy only and were regarded as controls, 30 others received 3 x 2 mg/day of an ergot alkaloid (co-dergocrine mesylate), and 30 others received 3 x 400 mg pentoxifylline (slow-release formulation)/day orally. Therapy was performed for eight weeks and CBF measured before start of treatment, after a four-week treatment period, and at the end of the study. CBF did not change significantly in the controlmore » group; both the pentoxifylline and the ergot alkaloid group presented with a significant increase in the CBF. This positive effect was significantly more pronounced in the pentoxifylline group and affected more ischemic than other brain tissues. In addition, symptoms like sleep disturbances, vertigo, and tinnitus improved significantly during the pentoxifylline observation period.« less

In vitro pathogenicity assay for the ergot fungus Claviceps purpurea.

PubMed

Scheffer, Jan; Tudzynski, Paul

2006-04-01

The pathogenic development of the biotrophic ergot fungus Claviceps purpurea is strictly limited to the ovary of grasses. Early colonization stages occur within a defined spatio-temporal course of events, including the directed growth to the vascular tissue for nutrient supply. To characterize mutant strains with putative defects in pathogenicity, the close observation of the infection pathway is therefore indispensable. Here, we describe the establishment of a new pathogenicity assay, based on the in vitro cultivation of isolated rye ovaries. The pathogenic development of a wild-type strain of C. purpurea was compared with the infection of mature rye flowers on whole plants. Up to the sixth day post inoculation, the route of infection within the isolated ovaries was maintained and temporally equal to that seen in mature flowers. Therefore, the in vitro pathogenicity assay is an effective alternative to the whole-plant infection tests, and suitable for detailed infection studies and screening high numbers of mutants for defects in early pathogenesis.

Occurrence of Ochratoxins, Fumonisin B2 , Aflatoxins (B1 and B2 ), and Other Secondary Fungal Metabolites in Dried Date Palm Fruits from Egypt: A Mini-Survey.

PubMed

Abdallah, Mohamed F; Krska, Rudolf; Sulyok, Michael

2018-02-01

This study was conducted to investigate the natural co-occurrence of 295 fungal and bacterial metabolites in 28 samples of dried date palm fruits collected from different shops distributed in Assiut Governorate, Upper Egypt in 2016. Extraction and quantification of the target analytes were done using the "dilute and shoot" approach followed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. In total, 30 toxic fungal metabolites were detected. Among these metabolites, 4 types of ochratoxins including ochratoxin type A and B were quantified in 3 samples (11%) with a contamination range from 1.48 to 6070 μg/kg for ochratoxin A and from 0.28 to 692 μg/kg for ochratoxin B. In addition, fumonisin B 2 was observed in 2 (7%) samples with contamination levels ranging from 4.99 to 16.2 μg/kg. The simultaneous detection of fumonisin B 2 in the same contaminated samples with ochratoxins indicates the fungal attack by Aspergillus niger species during storage. Only 1 sample was contaminated with aflatoxin B 1 (14.4 μg/kg) and B 2 (2.44 μg/kg). The highest maximum concentration (90400 μg/kg) was for kojic acid that contaminated 43% of the samples. To the best of the authors' knowledge, this is the first report of the natural co-occurrence of fumonisin B 2 and ochratoxin A and B in addition to a wide range of other fungal metabolites in date palm fruits. Mycotoxins are secondary metabolites produced by different fungi. These metabolites pose a potential risk on human health since they contaminate many food commodities. Among these, date palm fruits which are an integral part of diet in several countries. Therefore, detection of mycotoxins is a prerequisite to insure the safety of food. Here, different types of mycotoxins have been detected in levels that may have health hazard. © 2018 Institute of Food Technologists®.

Effect of ergot alkaloids on serum prolactin in non-psychotic organic brain syndrome of the elderly.

PubMed

Gross, R J; Eisdorfer, C E; Schiller, H S; Cox, G

1979-08-01

A compound of ergot alkaloids (Hydergine-Sandoz) or placebo was given to sixty elderly nursing home patients with non-psychotic organic brain syndrome. Subsequent to 12 weeks of treatment, the mean serum prolactin of the drug was significantly lower than that of the placebo group. No correlation was found between changes in prolactin and changes in behavior using Sandoz Assessment of Clinical Status Rating Form-Geriatric [SCAG]. It may be that a higher dose of Hydergine with an accompanying greater drop in prolactin would be required to observe this effect.

Biodegradation of Ochratoxin A by Aspergillus tubingensis Isolated from Meju.

PubMed

Cho, Sung Min; Jeong, Seong Eun; Lee, Kyu Ri; Sudhani, Hemanth P K; Kim, Myunghee; Hong, Sung-Yong; Chung, Soo Hyun

2016-10-28

Ochratoxin A (OTA), a mycotoxin, contaminates agricultural products and poses a serious threat to public health worldwide. Microbiological methods are known to be a promising approach for OTA biodegradation because physical and chemical methods have practical limitations. In the present study, a total of 130 fungal isolates obtained from 65 traditional Korean meju (a fermented starter for fermentation of soybeans) samples were examined for OTA-biodegradation activity using thin-layer chromatography. Two fungal isolates were selected for OTA-biodegradation activity and were identified as Aspergillus tubingensis M036 and M074 through sequence analysis of the beta-tubulin gene. After culturing both A. tubingensis isolates in Soytone-Czapek medium containing OTA (40 ng/ml), OTA-biodegradation activity was analyzed using high-performance liquid chromatography (HPLC). Both A. tubingensis strains degraded OTA by more than 95.0% after 14 days, and the HPLC analysis showed that the OTA biodegradation by the A. tubingensis strains led to the production of ochratoxin α, which is much less toxic than OTA. Moreover, crude enzymes from the cultures of A. tubingensis M036 and M074 led to OTA biodegradation of 97.5% and 91.3% at pH 5, and 80.3% and 75.3% at pH 7, respectively, in a buffer solution containing OTA (40 ng/ml) after 24 h. In addition, the OTA-biodegrading fungi did not exhibit OTA production activity. Our data suggest that A. tubingensis isolates and their enzymes have the potential for practical application to reduce levels of OTA in food and feed.

Toxic effects, metabolism, and carry-over of ergot alkaloids in laying hens, with a special focus on changes of the alkaloid isomeric ratio in feed caused by hydrothermal treatment.

PubMed

Dänicke, Sven

2016-02-01

Ergot alkaloids (EA) are mycotoxins formed by Claviceps purpurea. Due to the large variation in EA content, the mass proportion of ergot (hardened sclerotia) in animal diets is not suited to establish safe levels of EA. Therefore, the aim of the present study was to examine the dose-dependent effects of dietary EA on laying hens. Ergoty rye or ergot-free rye (control diet) was included in the diets either untreated or after hydrothermal treatment ("expansion"). The total EA levels in five different diets containing 0-3% of untreated or expanded rye were 0.1-14.56 mg/kg (untreated rye) and 0.08-13.03 mg/kg (expanded rye). The average EA reduction amounted to 11% due to expanding. The proportions of the sum of all -inine isomers however were consistently higher (19.5-48.4%) compared to the sum of their -ine isomer counterparts which decreased at the same time. Most of the laying performance and reproductive traits were significantly compromised during the test period between weeks 22 and 42 of age when the diet with the highest EA content was fed. Toxic effects were less pronounced due to expanding. Relative weights of liver, proventriculus, and gizzard as well as the aspartate aminotransferase activity, the antibody titers to Newcastle disease virus, albumin, and total bilirubin concentrations were all significantly increased in hens fed at the highest dietary ergot level whereby expanding additionally modified the albumin and total bilirubin responses. No carry-over of EA into egg yolk and albumen, blood, liver, and breast muscle was found, but bile contained quantifiable levels of ergometrine and ergometrinine. Biological recovery of ingested individual alkaloids with the excreta varied from 2 to 22% and was strongly positive linearly related to the octanol to water partition coefficient (logkOW). This suggests the lipophilicity of alkaloids as a factor influencing their metabolism and elimination. Based on the overall results of this study, a lowest observed

Ochratoxin A in Portugal: A Review to Assess Human Exposure

PubMed Central

Duarte, Sofia C.; Pena, Angelina; Lino, Celeste M.

2010-01-01

In Portugal, the climate, dietary habits, and food contamination levels present the characteristics for higher population susceptibility to ochratoxin A (OTA), one of the known mycotoxins with the greatest public health and agro-economic importance. In this review, following a brief historical insight on OTA research, a summary of the available data on OTA occurrence in food (cereals, bread, wine, meat) and biological fluids (blood, urine) is made. With this data, an estimation of intake is made to ascertain and update the risk exposure estimation of the Portuguese population, in comparison to previous studies and other populations. PMID:22069635

Ergot alkaloids from endophyte-infected tall fescue decrease reticulo-ruminal epithelial blood flow and volatile fatty acid absorption from a washed reticulorumen

USDA-ARS?s Scientific Manuscript database

An experiment was conducted to determine if ergot alkaloids affect blood flow to the absorptive surface of the rumen. Steers (n = 8) were pair-fed alfalfa cubes and received ground endophyte-infected tall fescue seed (E+; 0.015 mg ergovaline•kg BW-1•d-1) or endophyte-free tall fescue seed (E-) via r...

Long-lasting ergot lipids as new biomarkers for assessing the presence of cereals and cereal products in archaeological vessels.

PubMed

Lucejko, Jeannette J; La Nasa, Jacopo; Porta, Francesca; Vanzetti, Alessandro; Tanda, Giuseppa; Mangiaracina, Claudio Filippo; Corretti, Alessandro; Colombini, Maria Perla; Ribechini, Erika

2018-03-02

Cereals were very important in ancient diets, however evidence from archaeological sites of the vessels used for processing or storing cereals is comparatively rare. Micro-organisms, as well as chemical-physical effects can easily degrade cereals during the burial period. This can lead to a complete cereal decay and to serious difficulties in estimating the intensity of use of the cereals by ancient populations. Here, we present a novel biomarker approach entailing the detection of secondary lipid metabolites produced by ergot fungi (genus Claviceps), which are common cereal pests. The aim was to identify the original presence of Gramineae and to indirectly establish if vessels were used for cereal storage/processing. The fatty acid and TAG-estolide profiles of the remains from more than 30 archaeological vessels were investigated by gas chromatography/mass spectrometry (GC/MS) and high performance liquid chromatography/high resolution mass spectrometry (HPLC/ESI-Q-ToF). The detection of lipids derived from ergot in archaeological and historic contexts rests on its complex chemistry, providing a unique and relatively recalcitrant chemical signature for cereals. This research demonstrated that the combination of our innovative biomarker approach along with environmental and archaeological evidence can provide unprecedented insights into the incidence of cereals and related processing activities in ancient societies.

Application of upconversion luminescent-magnetic microbeads with weak background noise and facile separation in ochratoxin A detection

NASA Astrophysics Data System (ADS)

Liao, Zhenyu; Zhang, Ying; Su, Lin; Chang, Jin; Wang, Hanjie

2017-02-01

Ochratoxin A (OTA), the most harmful and abundant ochratoxin, is chemically stable and commonly existed in foodstuffs. In this work, upconversion luminescent-magnetic microbeads (UCLMMs) -based cytometric bead array for OTA detection with a less reagent consumption and high sensitivity has been established and optimized. In UCLMMs, upconversion nanocrystals (UCNs) for optical code present a weak background noise and no spectral cross talk between the encoding signals and target labels under two excitation conditions to improve detection sensitivity. While the superparamagnetic Fe3O4 nanoparticles (Fe3O4 NPs) aim for rapid analysis. The results show that the developed method has a sensitivity of 9.553 ppt below HPLC with a 50-μL sample and can be completed in <2 h with good accuracy and high reproducibility. Therefore, different colors of UCLMMs will become a promising assay platform for multiple mycotoxins after further improvement.

Trichothecene Genotype Composition of Fusarium graminearum Not Differentiated Among Isolates from Maize Stubble, Maize Ears, Wheat Spikes, and the Atmosphere in New York.

PubMed

Kuhnem, Paulo R; Spolti, Pierri; Del Ponte, Emerson M; Cummings, Jaime A; Bergstrom, Gary C

2015-05-01

In order to test the hypothesis that the trichothecene genotype composition of local populations of Fusarium graminearum is structured by specific habitats, a collection of 1,407 isolates was obtained from overwintered maize stubble, mature maize ears and wheat spikes, and the atmosphere 1.5 m aboveground during the flowering stage of these crops. These isolates were sampled at three diverse agricultural locations in New York State: namely, Aurora (sampled in 2012 and 2013) in central New York, Belmont (sampled in 2013) in southwestern New York, and Willsboro (sampled in 2013) in northeastern New York. Approximately 100 isolates of F. graminearum from each habitat were collected within a 10-mile2 area in each location. Polymerase chain reaction assays were used to identify three main B-trichothecene genotypes--3-acetyldeoxynivalenol (3-ADON), 15-ADON, or nivalenol (NIV)--based on amplification of portions of Tri3 and Tri12 genes. All but the NIV genotype were detected. The 15-ADON genotype predominated in most locations; frequencies were 92% (652/709) at Aurora, 78% (332/379) at Belmont, and 53% (167/319) at Willsboro. Frequencies of any genotype did not differ in general among the four habits in each location. An exception was in Aurora 2012, where only 5 in 24 3-ADON isolates were found in samplings from the air and grains of both crops. As viewed by the composition of trichothecene genotypes, local populations of F. graminearum appear not to be structured by these four habitats inclusive of pathogenic and saprophytic phases of the fungus life cycle. The similar frequency of 3-ADON and 15-ADON in eastern New York (Willsboro), which is less than 400 km away from the Aurora sampling location in the central area of the state, suggests that regional populations may be differentiated based on selection associated with climatic or landscape features not currently identified.

Functionalized Ergot-alkaloids as potential dopamine D3 receptor agonists for treatment of schizophrenia

NASA Astrophysics Data System (ADS)

Ivanova, Bojidarka; Spiteller, Michael

2012-12-01

The relationship between the molecular structure and physical properties of functionalized naturally occurred Ergot-alkaloids as potential dopamine D3 receptor agonists is presented. The molecular modeling of the ergoline-skeleton is based on the comprehensive theoretical study of the binding affinity of the isolated chemicals towards the active sites of the D3 sub-type receptor (D3R) loops. The studied proton accepting ability under physiological conditions allows classifying four types of monocationics, characterizing with the different binding modes to D3R involving selected amino acid residues to the active sites. These results marked the pharmaceutical potential and clinical usage of the reported compounds as antipsychotic drugs for Schizophrenia treatment, since they allowed evaluating the highlights of the different hypothesizes of the biochemical causes the illness. The applied complex approach for theoretical and experimental elucidation, including quantum chemistry method, electrospray ionization (ESI) and matrix assisted laser desorption/ionization (MALDI) mass spectrometric (MS) methods, nuclear magnetic resonance and vibrational IR and Raman spectroscopy on the isolated fifteen novel derivatives (1)-(15) and their different protonated forms (1a)-(15a) evidenced a strong dependence of molecular conformation, physical properties and binding affinity. Thus, the semi-synthetic functionalization of the naturally occurred products (NPs), provided significant possibilities to further molecular drugs-design and development of novel derivatives with wanted biological function, using the established profile of selected classes/families of NPs. The work described chiefly the non-linear (NL) approach for the interpretation of the mass chromatograms on the performed hybrid high performance liquid chromatography (HPLC) tandem MS/MS and MS/MS/MS experiments, discussing the merits and great diversity of instrumentation flexibility, thus achieving fundamental

Determination of aflatoxins and ochratoxin A in high-sugar-content traditional Turkish foods by affinity column cleanup and LC fluorescence detection.

PubMed

Senyuva, Hamide Z; Cimen, Dilek; Gilbert, John

2009-01-01

The effectiveness of an affinity column cleanup procedure followed by LC with fluorescence detection was established for the determination of aflatoxins and ochratoxin A in high-sugar-content traditional Turkish foods. Traditional foods, such as baklava (finely layered pastry filled with nuts and steeped in syrup), halvah (containing sesame paste and pistachios), cevizli sucuk (a confection made of grape juice boiled and dried on strings of nuts), Turkish delight (containing hazelnuts, pistachios, or walnuts), and pişmaniye (candy made of sugar, butter, and flour), were tested, and the performance of the method was established with spiked samples. To examine the robustness of the methodology, baklava was prepared from raw materials and spiked at the initial stage of dry ingredients and through subsequent stages of preparation of dough, after cooking, and after addition of syrup and nuts. For all products, the analytical method required grinding the composite foodstuff under liquid nitrogen to form a fine powder, which was then thoroughly mixed before subsampling. After vortex extraction into methanol-water (aflatoxins) and aqueous sodium bicarbonate (ochratoxin A), the sample was filtered, diluted with phosphate-buffered saline, and then passed through either an aflatoxin or ochratoxin A affinity column before HPLC analysis with fluorescence detection (using post-column bromination for the aflatoxins). In all the traditional Turkish products, the recovery of aflatoxin B1 ranged from 77 to 98%, and LODs were <0.1 microg/kg. For ochratoxin A, the recoveries were from 88 to 93% and LODs were similarly <0.1 microLg/kg. Despite the complex nature of these traditional Turkish foods, which frequently contain products from sugar caramelization, there was no evidence of any interfering co-extractives, and the method has proved to be robust enough to be used for food control purposes.

Surface plasmon resonance biosensor for the detection of ochratoxin A in cereals and beverages.

PubMed

Yuan, Jing; Deng, Dawei; Lauren, Denis R; Aguilar, Marie-Isabel; Wu, Yinqiu

2009-12-10

Ochratoxins are a group of mycotoxins produced as secondary metabolites by fungi which contaminate a large variety of food and feed commodities. Due to their teratogenic and carcinogenic properties, ochratoxins present a serious hazard to human and animal health. There is an increasing need to establish a simple sensitive method to detect these toxins. Here we report a rapid and highly sensitive surface plasmon resonance (SPR) assay of ochratoxin A (OTA) using Au nanoparticles for signal enhancement on a mixed self-assembled monolayer (mSAM) surface. A competitive immunoassay format was used for the development of the OTA immunoassay, which is based on the immobilization of target OTA through its ovalbumin (OVA) conjugate with a polyethylene glycol (PEG) linker. The new OTA conjugate (OTA-PEG-OVA) showed remarkably enhanced performance characteristics compared with those based on the immobilization of a commercial bovine serum albumin BSA-OTA conjugate without a PEG linker. Although OTA concentrations as low as 1.5 ng mL(-1) could be directly detected on this surface, the limit of detection (LOD) can be dramatically improved to 0.042 ng mL(-1) for OTA by applying large gold nanoparticles (40 nm) for signal enhancement. Various chemical conditions to minimize the influence of the food matrix on assay performance were also investigated. Grain samples were simply extracted with 50% methanol and liquid samples treated with poly(vinylpyrrolidone) (PVP) (3 or 5%), without any sample clean-up or pre-concentration step prior to analysis. The LODs for OTA in oats and corn were 0.3 and 0.5 ng g(-1), respectively, while in wine and other beverages, LODs ranged from 0.058 to 0.4 ng mL(-1). No cross-reactivity was observed with three other common mycotoxins. In addition, the mSAM/OTA-PEG-OVA surface exhibited high stability with over 600 binding/regeneration cycles. This approach with simple sample preparation provides a powerful tool for the rapid and sensitive quantitative

[Simultaneous determination of ochratoxin A, B and citrinin in foods by HPLC-FL and LC/MS/MS].

PubMed

Tabata, Setsuko; Iida, Kenji; Kimura, Keisuke; Iwasaki, Yumiko; Nakazato, Mitsuo; Kamata, Kunihiro; Hirokado, Masako

2008-04-01

Methods using high-performance liquid chromatography with fluorescence detection (HPLC-FL) and using liquid chromatography with tandem mass spectrometry (LC/MS/MS) were developed for simultaneous determination of ochratoxin A (OTA), ochratoxin B (OTB) and citrinin (CIT) in cereal, fruit, and coffee products. The samples were extracted with ethyl acetate under an acidic condition, and then cleaned up with liquid-liquid separation. The test solutions were analyzed by reverse-phase HPLC-FL and LC/MS/MS. Mass spectral acquisition was performed in positive ion mode by applying multiple reaction monitoring. The performances of both detectors were almost equivalent. The recoveries of OTA and OTB were 87-111%, and that of CIT were 70-88%. The limits of quantification (S/N> or =10) of OTA, OTB and CIT was 0.1 mug/kg or less. These methods were considered to be useful for the determination of the three mycotoxins at low levels (0.1 microg/kg).

Peptide YY3–36 and 5-Hydroxytryptamine Mediate Emesis Induction by Trichothecene Deoxynivalenol (Vomitoxin)

PubMed Central

Pestka, James J.

2013-01-01

Deoxynivalenol (DON, vomitoxin), a trichothecene mycotoxin produced by Fusarium sp. that frequently occurs in cereal grains, has been associated with human and animal food poisoning. Although a common hallmark of DON-induced toxicity is the rapid onset of emesis, the mechanisms for this adverse effect are not fully understood. Recently, our laboratory has demonstrated that the mink (Neovison vison) is a suitable small animal model for investigating trichothecene-induced emesis. The goal of this study was to use this model to determine the roles of two gut satiety hormones, peptide YY3–36 (PYY3–36) and cholecystokinin (CCK), and the neurotransmitter 5-hydroxytryptamine (5-HT) in DON-induced emesis. Following ip exposure to DON at 0.1 and 0.25mg/kg bw, emesis induction ensued within 15–30min and then persisted up to 120min. Plasma DON measurement revealed that this emesis period correlated with the rapid distribution and clearance of the toxin. Significant elevations in both plasma PYY3–36 (30–60min) and 5-HT (60min) but not CCK were observed during emesis. Pretreatment with the neuropeptide Y2 receptor antagonist JNJ-31020028 attenuated DON- and PYY-induced emesis, whereas the CCK1 receptor antagonist devezapide did not alter DON’s emetic effects. The 5-HT3 receptor antagonist granisetron completely suppressed induction of vomiting by DON and the 5-HT inducer cisplatin. Granisetron pretreatment also partially blocked PYY3–36-induced emesis, suggesting a potential upstream role for this gut satiety hormone in 5-HT release. Taken together, the results suggest that both PYY3–36 and 5-HT play contributory roles in DON-induced emesis. PMID:23457120

Development of ochratoxin a in cereal by chemiluminescence enzyme immunoassay

NASA Astrophysics Data System (ADS)

Li, Min; Liu, Renrong; Zhu, Lixin; Chen, Zhenzhen

2017-11-01

A rapid, simple and sensitive chemiluminescence enzyme immunoassay method (CLEIA) was established to detect ochratoxin A (OTA) in cereal. Optimal conditions including antibody dilution ratio and enzyme conjugate, ionic strength, pH value and organic solvent. Established indirect competition inhibition curve to determine the linear working range, detection limit and recovery rate. Results: The 50% inhibitory concentration and the detection limit of the CLEIA were78.8pg/mL and 14.86 pg/mL, respectively, with a linear range of 0.015-0.4ng/mL. At 1∼4μpg/kg fortified levels in wheat, mean recoveries ranged from 67.47% to100.35%.

Analysis of trichothecene mycotoxins in contaminated grains by gas chromatography/matrix isolation/Fourier transform infrared spectroscopy and gas chromatography/mass spectrometry.

PubMed

Mossoba, M M; Adams, S; Roach, J A; Trucksess, M W

1996-01-01

Gas chromatography/matrix isolation/Fourier transform infrared (GC/MI/FTIR) spectroscopy and GC/mass spectrometry (MS) were used to confirm the identities of trimethylsilyl (TMS) derivatives of trichothecene mycotoxins in naturally contaminated grains. Infrared spectral bands observed in the fingerprint region were unique for 10 trichothecene standards. Characteristic absorption bands were observed for the ester (near 1750 cm-1) and ketone (near 1700 cm-1) carbonyl stretching vibrations, the acetate CH3 symmetric bend (1370 cm-1), the epoxide ring (1262 cm-1), the trimethylsilyl CH3 in-plane deformation (1253 cm-1), the ester (O)C-O asymmetric stretching vibration (near 1244 cm-1), and several other bands including intense features due to the TMS function. Infrared bands observed under cryogenic matrix isolation conditions were compared with those found at room temperature in a potassium bromide matrix for 5 of these standards. Identities of deoxynivalenol (DON) from barley and mixed feed, nivalenol from wheat and barley, and DON and fusarenon-x from sweet corn were confirmed by comparison of their infrared spectral bands with those of standards. The identity of DON in the same test samples of sweet corn was confirmed further by GC/MS. GC/MS was also used to quantitate the levels of DON (67-455 ppm) in sweet corn test samples.

Natural occurrence of ochratoxin A contamination in commercial black and white pepper products.

PubMed

Jalili, M; Jinap, S; Radu, S

2010-10-01

The concentration of ochratoxin A (OTA) in 120 commercial pepper (84 pre-packed and 36 bulk samples), which consist of local and imported white and black pepper in powder and seed form in Malaysia were determined. The objective of the study was to investigate and compare OTA concentration in black pepper and white pepper being commercialized in Malaysia. Determination method was based on HPLC with fluorescence detection coupled with immunoaffinity column clean-up step. Mobile phase consisted of acetonitrile-water-acetic acid (49.5:49.5:1.0, v/v/v), and flow rate was 1 ml/min. The LOD was 0.02 ng/g, and the average recovery values of OTA ranged from 79.5 to 92.0% in black pepper and 81.2-90.3% in white pepper. A total of 57 samples (47.5%) were contaminated with OTA ranging from 0.15 to 13.58 ng/g. The results showed that there was a significant difference between type of pepper and brands. OTA concentration in black pepper was significantly higher than white pepper (p < 0.05). The highest concentration of ochratoxin, 13.58 ng/g, was detected in a sample of black pepper seed followed by 12.64 ng/g in a sample of black pepper powder, both were bulk samples purchased from open market.

Detection of Ochratoxin a Using Molecular Beacons and Real-Time PCR Thermal Cycler

PubMed Central

Sanzani, Simona Marianna; Reverberi, Massimo; Fanelli, Corrado; Ippolito, Antonio

2015-01-01

We developed a simple and cheap assay for quantitatively detecting ochratoxin A (OTA) in wine. A DNA aptamer available in literature was used as recognition probe in its molecular beacon form, i.e., with a fluorescence-quenching pair at the stem ends. Our aptabeacon could adopt a conformation allowing OTA binding, causing a fluorescence rise due to the increased distance between fluorophore and quencher. We used real-time PCR equipment for capturing the signal. With this assay, under optimized conditions, the entire process can be completed within 1 h. In addition, the proposed system exhibited a good selectivity for OTA against other mycotoxins (ochratoxin B and aflatoxin M1) and limited interference from aflatoxin B1 and patulin. A wide linear detection range (0.2–2000 µM) was achieved, with LOD = 13 nM, r = 0.9952, and R2 = 0.9904. The aptabeacon was also applied to detect OTA in red wine spiked with the same dilution series. A linear correlation with a LOD = 19 nM, r = 0.9843, and R2 = 0.9708 was observed, with recoveries in the range 63%–105%. Intra- and inter-day assays confirmed its reproducibility. The proposed biosensor, although still being finalized, might significantly facilitate the quantitative detection of OTA in wine samples, thus improving their quality control from a food safety perspective. PMID:25760080

Titanium Dioxide Nanoparticles (TiO₂) Quenching Based Aptasensing Platform: Application to Ochratoxin A Detection.

PubMed

Sharma, Atul; Hayat, Akhtar; Mishra, Rupesh K; Catanante, Gaëlle; Bhand, Sunil; Marty, Jean Louis

2015-09-22

We demonstrate for the first time, the development of titanium dioxide nanoparticles (TiO₂) quenching based aptasensing platform for detection of target molecules. TiO₂ quench the fluorescence of FAM-labeled aptamer (fluorescein labeled aptamer) upon the non-covalent adsorption of fluorescent labeled aptamer on TiO₂ surface. When OTA interacts with the aptamer, it induced aptamer G-quadruplex complex formation, weakens the interaction between FAM-labeled aptamer and TiO₂, resulting in fluorescence recovery. As a proof of concept, an assay was employed for detection of Ochratoxin A (OTA). At optimized experimental condition, the obtained limit of detection (LOD) was 1.5 nM with a good linearity in the range 1.5 nM to 1.0 µM for OTA. The obtained results showed the high selectivity of assay towards OTA without interference to structurally similar analogue Ochratoxin B (OTB). The developed aptamer assay was evaluated for detection of OTA in beer sample and recoveries were recorded in the range from 94.30%-99.20%. Analytical figures of the merits of the developed aptasensing platform confirmed its applicability to real samples analysis. However, this is a generic aptasensing platform and can be extended for detection of other toxins or target analyte.

Fluorometric aptamer based assay for ochratoxin A based on the use of exonuclease III.

PubMed

Liu, Renjie; Wu, Hua; Lv, Lei; Kang, Xiaojiao; Cui, Chengbi; Feng, Jin; Guo, Zhijun

2018-04-14

This study describes an aptamer based assay for the mycotoxin ochratoxin A (OTA). The method is based on the use of an OTA-specific aptamer, exonuclease (Exo) III, SYBR Gold as a fluorescent probe, and a complementary strand that specifically combines with the aptamer. In the presence of OTA, the aptamer and OTA hybridize, thereby resulting in the formation of ssDNA, which is not digested by Exo III. Intense fluorescence is observed after addition of SYBR Gold (best measured at excitation/emission wavelengths of 495/540 nm). Fluorescence increases linearly with the log of the OTA concentration in the range from 8 to 1000 ng·mL -1 . The detection limit is 4.7 ng·mL -1 . The assay was applied to the determination of OTA in diluted [2%(v/v)] red wine, and recoveries and RSDs ranged between 93.5% and 113.8%, and between 3.2% and 5.7%, respectively. Graphical abstract In the presence of ochratoxin A (OTA), specific combinations of aptamer and OTA may occur and result in DNA double strands being untied, which avoids being digested by Exo III. Intense fluorescence is observed after SYBR Gold addition.

Immunotoxicity of ochratoxin A and aflatoxin B1 in combination is associated with the nuclear factor kappa B signaling pathway in 3D4/21 cells.

PubMed

Hou, Lili; Gan, Fang; Zhou, Xuan; Zhou, Yajiao; Qian, Gang; Liu, Zixuan; Huang, Kehe

2018-05-01

The co-contamination of cereals, grains, crops, and animal feeds by mycotoxins is a universal problem. Humans and animals are exposed to several mycotoxins simultaneously as evidenced by extensive studies on this topic. Yet, most studies have addressed the effects of mycotoxins individually. Aflatoxin B1 and ochratoxin A can induce immunotoxicity. However, it remains unclear whether a combination of these mycotoxins aggravates immunotoxicity and the potential mechanism underlying this effect. In this study, we used the cell line 3D4/21, swine alveolus macrophages and innate immune cell. The results showed that the percentage of cell inhibition, annexin V/PI-positive rates, and the expression of pro-inflammatory cytokines (tumor necrosis factor alpha and interleukin-6) significantly increased and the release of lactate dehydrogenase and phagocytotic index were significantly decreased at different concentrations of aflatoxin B1 and ochratoxin A combination when compared with control. The combination of aflatoxin B1 and ochratoxin A significantly decreased the production of GSH and increased reactive oxygen species level. However, N-acetylcysteine suppressed the oxidative stress and alleviated the immunotoxicity induced by the combination. The combination of aflatoxin B1 and ochratoxin A markedly enhanced the degradation of IκBa, the phosphorylation of nuclear factor kappa B (p65), and the translocation of activated nuclear factor kappa B (NF-κB) into the nuclei as demonstrated by western blotting and confocal laser scanning microscopy. These effects could be reversed by BAY 11-7082, a specific inhibitor of NF-κB. Taken together, a combination of aflatoxin B1 and ochratoxin A could aggravate immunotoxicity by activating the NF-κB signaling pathway. Copyright © 2018 Elsevier Ltd. All rights reserved.

Effects of macrocyclic trichothecene mycotoxins on the murine immune system

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hughes, B.J.

1988-01-01

The macrocyclic trichothecenes are a unique group of toxins which have some antileukemic properties. In the first study, verrucarin A and roridin A were examined. Both mycotoxins were administered intraperitoneally at an equitoxic dose of 0.35 mg/kg to CD-1 mice. Lymphocyte proliferation was studied after animals were dosed with verrucarin A. After day 2, no differences in {sup 3}H-thymidine incorporation were observed using concanavalin A (Con A), phytohemagglutinin (PHA), pokeweed mitogen (PWM), or lipopolysaccharide (LPS). On day 4, DNA synthesis induced by Con A, PHA, and PWM increased significantly. On day 7, PHA stimulation increased above controls while Con A,more » PWM, and LPS responses were not significantly different. In contrast, roridin A decreased PHA stimulation only on day 7. In the second study the mycotoxins roritoxin B, myrotoxin B, roridin A, verrucarin A, 16-hydroxyverrucarin A, verrucarin J, baccharinoid B12, roridin D, roridin E, baccharinoid B4, and baccharinoid B5 were investigated. In the third study lymphocytes were cultured with each of the mycotoxins for 48 hr to assess their lethality.« less

Method for analysis dried vine fruits contaminated with ochratoxin A.

PubMed

Galvis-Sánchez, Andrea C; Barros, Antonio S; Delgadillo, Ivonne

2008-06-09

The EU maximum limit of 10 microg kg(-1) of OTA for dried vine fruits has been established since 2002 (European Commission, 2005). The presented work explore the capability of using Fourier infrared spectroscopy attenuated total reflection (FTIR-ATR) for the detection of ochratoxin A (OTA) in dried vine fruits in a range of concentration between 2 and 50 microg kg(-1) OTA. The method developed included a sample pretreatment using a C18 cartridge which was efficient for the isolation of the mycotoxin. The PLS1 analysis of the spectrum of sultanas spiked with different OTA concentrations showed a good correlation between the spectral data and reference concentration for OTA (R(2)=0.85).

Environment factors influence in vitro interspecific interactions between A. ochraceus and other maize spoilage fungi, growth and ochratoxin production.

PubMed

Lee, H B; Magan, N

1999-01-01

The effect of water availability (water activity, aw; 0.995-0.90) and temperature (18-30 degrees C) on in vitro interactions between an ochratoxin producing strain of Aspergillus ochraceus and six other spoilage fungi was assessed in dual culture experiments on a maize meal-based agar medium. In primary resource capture of nutrient substrate, A. ochraceus was dominant against many of the interacting species, being able to overgrow and replace A. candidus, and sometimes A. flavus and the Eurotium spp. regardless of aw or temperature. However, with freely available water (0.995 aw) A. alternata and A. niger were dominant, with mutual antagonism between A. ochraceus and A. flavus at 25-30 degrees C. In the driest conditions tested (0.90 aw) there was also mutual antagonism between A. ochraceus and the two Eurotium spp. Overall, under all conditions tested the Index of Dominance for A. ochraceus was much higher than for other competing species combined suggesting that A. ochraceus was a good competitive colonist able to replace a number of other species. However, the growth rate of A. ochraceus was modified and decreased by the interaction with competitors. Interaction between A. ochraceus and species such as A. alternata (18 degrees C/0.995) and Eurotium spp. (0.995-0.95 and 25-30 degrees C) resulted in a significant stimulation of ochratoxin production. The results are discussed in relation to the effect that environmental factors have on the possible competitiveness of A. ochraceus in the maize grain ecosystem and the role of ochratoxin in niche exclusion of competitors.

Study on the presence of ochratoxin α in cultures of ochratoxigenic and non- ochratoxigenic strains of Aspergillus carbonarius.

PubMed

Bragulat, M Rosa; Eustaquio, Alba; Cabañes, F Javier

2017-01-01

Ochratoxin A (OTA) is a potent nephrotoxin and carcinogen which is found in a wide variety of common foods and beverages and it is produced by several species of Aspergillus and Penicillium. Ochratoxin α (OTα), a major metabolite of OTA, has also been reported to occur in cultures of OTA-producing species. However there is some controversial about the participation of OTα in the biosynthesis of OTA, mainly because its biosynthesis pathway has not yet been completely characterized. Aspergillus carbonarius is the main responsible source of ochratoxin A (OTA) in food commodities such as wine, grapes or dried vine fruits from main viticultural regions worldwide. However, little is known about the presence of OTα in isolates of A. carbonarius. In this study we evaluated the effects of temperature and incubation time on OTα production by both OTA and non-OTA-producing strains of A. carbonarius. OTA and OTα were detected on the basis of HPLC fluorometric response compared with that of their standards and confirmed by HPLC-MS in selected samples. The non-OTA-producing strains did produce neither OTA nor OTα at any of the conditions tested. The OTA-producing strains studied were able to produce both OTA and OTα in most of the conditions tested. In general, higher amounts of OTA than OTα were produced, but a positive correlation in the production of these two metabolites was detected. The lack of production of both OTA and OTα in the non-OTA-producing strains could be caused by the presence of silent genes or by mutations in functional or regulatory genes involved in OTA production.

Study on the presence of ochratoxin α in cultures of ochratoxigenic and non- ochratoxigenic strains of Aspergillus carbonarius

PubMed Central

Bragulat, M. Rosa; Eustaquio, Alba

2017-01-01

Ochratoxin A (OTA) is a potent nephrotoxin and carcinogen which is found in a wide variety of common foods and beverages and it is produced by several species of Aspergillus and Penicillium. Ochratoxin α (OTα), a major metabolite of OTA, has also been reported to occur in cultures of OTA-producing species. However there is some controversial about the participation of OTα in the biosynthesis of OTA, mainly because its biosynthesis pathway has not yet been completely characterized. Aspergillus carbonarius is the main responsible source of ochratoxin A (OTA) in food commodities such as wine, grapes or dried vine fruits from main viticultural regions worldwide. However, little is known about the presence of OTα in isolates of A. carbonarius. In this study we evaluated the effects of temperature and incubation time on OTα production by both OTA and non-OTA-producing strains of A. carbonarius. OTA and OTα were detected on the basis of HPLC fluorometric response compared with that of their standards and confirmed by HPLC-MS in selected samples. The non-OTA-producing strains did produce neither OTA nor OTα at any of the conditions tested. The OTA-producing strains studied were able to produce both OTA and OTα in most of the conditions tested. In general, higher amounts of OTA than OTα were produced, but a positive correlation in the production of these two metabolites was detected. The lack of production of both OTA and OTα in the non-OTA-producing strains could be caused by the presence of silent genes or by mutations in functional or regulatory genes involved in OTA production. PMID:29016677

Natural occurrence of fumonisins and ochratoxin A in some herbs and spices commercialized in Poland analyzed by UPLC-MS/MS method.

PubMed

Waśkiewicz, Agnieszka; Beszterda, Monika; Bocianowski, Jan; Goliński, Piotr

2013-12-01

Unsanitary conditions during harvesting, drying, packing and storage stages in production and processing of spices and herbs could introduce mycotoxin contamination. The occurrence of ochratoxin A and fumonisins in popular spices and herbs was studied, using liquid chromatography-electrospray-mass spectrometry. Apart from mycotoxins, ergosterol as a factor indicating fungal development was also analysed. A total of 79 different samples commercialized in Poland were randomly purchased from popular markets were tested for mycotoxins. The frequency of samples with fumonisins was lower (31%) than ochratoxin A (49%). Free from mycotoxins were samples of bay leaf and white mustard. ERG content - in spice samples with high concentration level of mycotoxins - was also significantly higher than in samples with little to no mycotoxins. Copyright © 2013 Elsevier Ltd. All rights reserved.

Role of Glycols and Tweens in the Production of Ergot Alkaloids by Claviceps paspali

PubMed Central

Mizrahi, A.; Miller, G.

1969-01-01

Several glycols and Tweens markedly stimulated the production of ergot alkaloids in submerged cultures of Claviceps paspali. The role of these compounds was investigated in shake flasks and bench-scale fermentors. 2,3-Butanediol was not utilized by the fungus, and 1,2-propanediol-1-14C was not incorporated into the alkaloids. Glycols and Tweens lowered the surface tension of the basal medium and promoted the utilization of metabolites. In the presence of glycols and Tweens, an increased uptake of labeled sorbitol and succinic acid took place, whereas the specific radioactivity of the alkaloids was not affected. These results indicated that glycols and Tweens are not involved directly in the biosynthetic process; they apparently acted as surface-active agents, facilitating transport of metabolites into the cells. PMID:5776521

Variants of Aspergillus alutaceus var. alutaceus (formerly Aspergillus ochraceus) with altered ochratoxin a production

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chelack, W.S.; Borsa, J.; Szekely, J.G.

1991-09-01

The present studies, using Asperigillus alutaceus var. alutaceus Berkeley et Curtis (formerly A. ochraceus Wilhelm) NRRL 3174 along with three other wild-type strains, were undertaken in an attempt to understand the effects of irradiation and other treatments on mycotoxin production in grain. Bedford barley was inoculated with spores of NRRL 3174, gamma irradiated, and incubated at 28C and 25% moisture. After 10 days of incubation, two colony types, ocher (parental) and yellow (variant), were isolated from the grain. Further culturing of the yellow variant resulted in the spontaneous appearance of a white variant that exhibited greatly enhanced fluorescence under UVmore » light. In subsequent work, we have also isolated variants producing a soluble red pigment. In addition, in model experiments involving irradiation (1 kGy) of pure cultures, induction frequencies ranging between 2 and 4% (survival basis) were observed for the yellow and red variants. Inoculation of these variants into wheat and incubation for 14 days at 28C and 32% moisture resulted in ochratoxin A production in the relative amounts of 0.09:1:4.6:9.3 for the red, ocher (parental), yellow, and white variants, respectively. Additional characteristics of these isolates are described. Confirmation that the white high-ochratoxin-A-producing variants were derived from the parental strain was demonstrated by obtaining revertant sectors in monoclonal cultures of the variants.« less

Use of a nonhomologous end joining deficient strain (Deltaku70) of the ergot fungus Claviceps purpurea for identification of a nonribosomal peptide synthetase gene involved in ergotamine biosynthesis.

PubMed

Haarmann, Thomas; Lorenz, Nicole; Tudzynski, Paul

2008-01-01

The ergot fungus Claviceps purpurea uses mainly the nonhomologous-end-joining (NHEJ) system for integration of exogenous DNA, leading to a low frequency of homologous integration (1-2%). To improve gene targeting efficiency we deleted the C. purpurea ku70 gene in two different strains: the pathogenic strain 20.1 and the apathogenic, ergot alkaloid producing strain P1. The mutants were not impaired in vegetative and pathogenic development nor alkaloid production. Gene targeting efficiency was significantly increased (50-60%) in the Deltaku70 mutants. The P1 Deltaku70 strain (producing ergotamine and ergocryptine) was used for targeted deletion of lpsA1, one of the two trimodular NRPS genes present in the alkaloid gene cluster, encoding D-lysergyl peptide synthetases involved in formation of the tripeptide moiety of ergopeptines. Mutants lacking the lpsA1 gene were shown to be incapable of producing ergotamine but were still able to produce ergocryptine, proving that LpsA1 is involved in ergotamine biosynthesis.

Ochratoxin A Producing Species in the Genus Penicillium

PubMed Central

Cabañes, Francisco Javier; Bragulat, Maria Rosa; Castellá, Gemma

2010-01-01

Ochratoxin A (OTA) producing fungi are members of the genera Aspergillus and Penicillium. Nowadays, there are about 20 species accepted as OTA producers, which are distributed in three phylogenetically related but distinct groups of aspergilli of the subgenus Circumdati and only in two species of the subgenus Penicillium. At the moment, P. verrucosum and P. nordicum are the only OTA producing species accepted in the genus Penicillium. However, during the last century, OTA producers in this genus were classified as P. viridicatum for many years. At present, only some OTA producing species are known to be a potential source of OTA contamination of cereals and certain common foods and beverages such as bread, beer, coffee, dried fruits, grape juice and wine among others. Penicillium verrucosum is the major producer of OTA in cereals such as wheat and barley in temperate and cold climates. Penicillium verrucosum and P. nordicum can be recovered from some dry-cured meat products and some cheeses. PMID:22069629

Recent Advances for the Detection of Ochratoxin A.

PubMed

Ha, Tai Hwan

2015-12-04

Ochratoxin A (OTA) is one of the mycotoxins secreted by Aspersillus and Penicillium that can easily colonize various grains like coffee, peanut, rice, and maize. Since OTA is a chemically stable compound that can endure the physicochemical conditions of modern food processing, additional research efforts have been devoted to develop sensitive and cost-effective surveillance solutions. Although traditional chromatographic and immunoassays appear to be mature enough to attain sensitivity up to the regulation levels, alternative detection schemes are still being enthusiastically pursued in an attempt to meet the requirements of rapid and cost-effective detections. Herein, this review presents recent progresses in OTA detections with minimal instrumental usage, which have been facilitated by the development of OTA aptamers and by the innovations in functional nanomaterials. In addition to the introduction of aptamer-based OTA detection techniques, OTA-specific detection principles are also presented, which exclusively take advantage of the unique chemical structure and related physicochemical characteristics.

Deleterious Effects of Mycotoxin Combinations Involving Ochratoxin A

PubMed Central

Klarić, Maja Šegvić; Rašić, Dubravka; Peraica, Maja

2013-01-01

Ochratoxin A (OTA) is a nephrotoxic mycotoxin with carcinogenic properties. Its presence was detected in various foodstuffs all over the world but with significantly higher frequency and concentrations in areas with endemic nephropathy (EN). Even though food is often contaminated with more than one mycotoxin, earlier studies focused on the occurrence and toxicology of only OTA. Only a limited number of surveys showed that OTA co-occurs in food with mycotoxins (citrinin-CIT, penicilic acid, fumonisin B1-FB1, aflatoxins-AF) which exert nephrotoxic, carcinogenic or carcinogen-promoting activity. This review summarises the findings on OTA and its co-occurrence with the mentioned mycotoxins in food as well as experimental data on their combined toxicity. Most of the tested mycotoxin mixtures involving OTA produced additive or synergistic effects in experimental models suggesting that these combinations represent a significant health hazard. Special attention should be given to mixtures that include carcinogenic and cancer-promoting mycotoxins. PMID:24189375

Occurrence of ochratoxin a contamination and detection of ochratoxigenic aspergillus species in retail samples of dried fruits and nuts

USDA-ARS?s Scientific Manuscript database

Ochratoxin A (OTA) is a mycotoxin produced by several species of Aspergillus and Penicillium and is a potential contaminant of a wide variety of food products. To determine the incidence of OTA contamination in dried fruits and tree nuts, retail packaged and bulk raisins, dates, figs, prunes, almon...

Mass Spectral Investigations on Toxins. 7. Detection and Accurate Quantitation of Picogram Quantities of Macrocyclic Trichothecenes in Brazilian Plant Samples by Direct Chemical Ionization-Mass Spectrometer/Mass Spectrometer Techniques

DTIC Science & Technology

1987-09-01

trichothecenes are naturally occurring di. and triesters of unsubstituted and substituted verrucarols. 1 -" The diesters are termed as roridins, satratoxins, and...Satratoxins produced M- ions very efficiently despite the nature of the CI reagent gases.’ 6 The protonated molecules of satratoxins formed under these

Aflatoxins and ochratoxin A in different cocoa clones (Theobroma cacao L.) developed in the southern region of Bahia, Brazil.

PubMed

Maciel, Leonardo Fonseca; Felício, Ana Lúcia de Souza Madureira; Miranda, Lucas Caldeirão Rodrigues; Pires, Tassia Cavalcante; Bispo, Eliete da Silva; Hirooka, Elisa Yoko

2018-01-01

Brazil is the sixth largest producer of cocoa beans in the world, after Côte d'Ivoire, Ghana, Indonesia, Nigeria and Cameroon. The southern region of Bahia stands out as the country's largest producer, accounting for approximately 60% of production. Due to damage caused by infestation of the cocoa crop with the fungus Moniliophthora perniciosa, which causes 'witch's broom disease', research in cocoa beans has led to the cloning of species that are resistant to the disease; however, there is little information about the development of other fungal genera in these clones, such as Aspergillus, which do not represent a phytopathogenicity problem but can grow during the pre-processing of cocoa beans and produce mycotoxins. Thus, the aim of this work was to determine the presence of aflatoxin (AF) and ochratoxin A (OTA) in cocoa clones developed in Brazil. Aflatoxin and ochratoxin A contamination were determined in 130 samples from 13 cocoa clones grown in the south of Bahia by ultra-performance liquid chromatography with a fluorescence detector. The method was evaluated for limit of detection (LOD) (0.05-0.90 μg kg -1 ), limit of quantification (0.10-2.50 μg kg -1 ) and recovery (RSD) (89.40-95.80%) for AFB 1 , AFB 2 , AFG 1 , AFG 2 and OTA. Aflatoxin contamination was detected in 38% of the samples in the range of ochratoxin A was positive in 18% of the samples in the range of

Ochratoxin A Dietary Exposure of Ten Population Groups in the Czech Republic: Comparison with Data over the World

PubMed Central

Ostry, Vladimir; Malir, Frantisek; Dofkova, Marcela; Skarkova, Jarmila; Pfohl-Leszkowicz, Annie; Ruprich, Jiri

2015-01-01

Ochratoxin A is a nephrotoxic and renal carcinogenic mycotoxin and is a common contaminant of various food commodities. Eighty six kinds of foodstuffs (1032 food samples) were collected in 2011–2013. High-performance liquid chromatography with fluorescence detection was used for ochratoxin A determination. Limit of quantification of the method varied between 0.01–0.2 μg/kg depending on the food matrices. The most exposed population is children aged 4–6 years old. Globally for this group, the maximum ochratoxin A dietary exposure for “average consumer” was estimated at 3.3 ng/kg bw/day (lower bound, considering the analytical values below the limit of quantification as 0) and 3.9 ng/kg bw/day (middle bound, considering the analytical values below the limit of quantification as 1/2 limit of quantification). Important sources of exposure for this latter group include grain-based products, confectionery, meat products and fruit juice. The dietary intake for “high consumers” in the group 4–6 years old was estimated from grains and grain-based products at 19.8 ng/kg bw/day (middle bound), from tea at 12.0 ng/kg bw/day (middle bound) and from confectionery at 6.5 ng/kg bw/day (middle bound). For men aged 18–59 years old beer was the main contributor with an intake of 2.60 ng/kg bw/day (“high consumers”, middle bound). Tea and grain-based products were identified to be the main contributors for dietary exposure in women aged 18–59 years old. Coffee and wine were identified as a higher contributor of the OTA intake in the population group of women aged 18–59 years old compared to the other population groups. PMID:26378578

Ochratoxin A Dietary Exposure of Ten Population Groups in the Czech Republic: Comparison with Data over the World.

PubMed

Ostry, Vladimir; Malir, Frantisek; Dofkova, Marcela; Skarkova, Jarmila; Pfohl-Leszkowicz, Annie; Ruprich, Jiri

2015-09-10

Ochratoxin A is a nephrotoxic and renal carcinogenic mycotoxin and is a common contaminant of various food commodities. Eighty six kinds of foodstuffs (1032 food samples) were collected in 2011-2013. High-performance liquid chromatography with fluorescence detection was used for ochratoxin A determination. Limit of quantification of the method varied between 0.01-0.2 μg/kg depending on the food matrices. The most exposed population is children aged 4-6 years old. Globally for this group, the maximum ochratoxin A dietary exposure for "average consumer" was estimated at 3.3 ng/kg bw/day (lower bound, considering the analytical values below the limit of quantification as 0) and 3.9 ng/kg bw/day (middle bound, considering the analytical values below the limit of quantification as 1/2 limit of quantification). Important sources of exposure for this latter group include grain-based products, confectionery, meat products and fruit juice. The dietary intake for "high consumers" in the group 4-6 years old was estimated from grains and grain-based products at 19.8 ng/kg bw/day (middle bound), from tea at 12.0 ng/kg bw/day (middle bound) and from confectionery at 6.5 ng/kg bw/day (middle bound). For men aged 18-59 years old beer was the main contributor with an intake of 2.60 ng/kg bw/day ("high consumers", middle bound). Tea and grain-based products were identified to be the main contributors for dietary exposure in women aged 18-59 years old. Coffee and wine were identified as a higher contributor of the OTA intake in the population group of women aged 18-59 years old compared to the other population groups.

Regulation of IL-8 promoter activity by verrucarin A in human monocytic THP-1 cells.

PubMed

Liu, Jun; Simmons, Steve O; Pei, Ruoting

2014-01-01

Macrocyclic trichothecenes have been frequently detected in fungi in water-damaged buildings and exhibited higher toxicity than the well-studied trichothecenes; however, the mechanism underlying their toxicity has been poorly understood. In this study, transcriptional regulation of the cytokine interleukin (IL)-8 by a macrocyclic trichothecene, verrucarin A (VA), in human monocytic THP-1 cells is reported. Consistent with previous findings, VA was 100-fold more cytotoxic than deoxynivalenol (DON), while ochratoxin A (OA) was not cytotoxic. In cells transduced with the wild-type IL-8 promoter luciferase construct, VA induced a biphasic dose response composed of an upregulation of luciferase expression at low concentrations of 0.01-1 ng/ml and a downregulation at high levels of 10 ng/ml and higher. In contrast, DON induced a sigmoid-shaped dose response with the EC50 of 11.6 ng/ml, while OA did not markedly affect the IL-8 expression. When cells were transduced with IL-8 promoter with a mutation of transcription factor nuclear factor-κB (NF-κB)-binding site, VA (1 ng/ml), DON (1000 ng/ml), and tumor necrosis factor (TNF) α (20 ng/ml)-induced luciferase expression were impaired. In addition, the NF-κB inhibitor caffeic acid phenethyl ester inhibited VA-, DON-, and TNFα-induced luciferase expression. Mutation of the CCAAT/enhancer-binding protein (CEBP) β binding site of the IL-8 promoter affected only DON-, but not VA- and TNFα-induced luciferase expression. Taken together, these results suggested that VA activated IL-8 promoter via an NF-κB-dependent, but not CEBPβ-dependent, pathway in human monocytes.

Real-time PCR detection of toxigenic Fusarium in airborne and settled grain dust and associations with trichothecene mycotoxins.

PubMed

Halstensen, Anne Straumfors; Nordby, Karl-Christian; Eduard, Wijnand; Klemsdal, Sonja Sletner

2006-12-01

Inhalation of immunomodulating mycotoxins produced by Fusarium spp. that are commonly found in grain dust may imply health risks for grain farmers. Airborne Fusarium and mycotoxin exposure levels are mainly unknown due to difficulties in identifying Fusarium and mycotoxins in personal aerosol samples. We used a novel real-time PCR method to quantify the fungal trichodiene synthase gene (tri5) and DNA specific to F. langsethiae and F. avenaceum in airborne and settled grain dust, determined the personal inhalant exposure level to toxigenic Fusarium during various activities, and evaluated whether quantitative measurements of Fusarium-DNA could predict trichothecene levels in grain dust. Airborne Fusarium-DNA was detected in personal samples even from short tasks (10-60 min). The median Fusarium-DNA level was significantly higher in settled than in airborne grain dust (p < 0.001), and only the F. langsethiae-DNA levels correlated significantly in settled and airborne dust (r(s) = 0.20, p = 0.003). Both F. langsethiae-DNA and tri5-DNA were associated with HT-2 and T-2 toxins (r(s) = 0.24-0.71, p < 0.05 to p < 00.01) in settled dust, and could thus be suitable as indicators for HT-2 and T-2. The median personal inhalant exposure to specific toxigenic Fusarium spp. was less than 1 genome m(-3), but the exposure ranged from 0-10(5) genomes m(-3). This study is the first to apply real-time PCR on personal samples of inhalable grain dust for the quantification of tri5 and species-specific Fusarium-DNA, which may have potential for risk assessments of inhaled trichothecenes.

The Effects of Ochratoxin A on the Immune Response of Swiss Mice

PubMed Central

Prior, M.G.; Sisodia, C.S.

1982-01-01

The acute intraperitoneal toxicity of ochratoxin A (OA) for adult female Swiss mice is presented. The seven-day LD50 was calculated to be 48 ± 3.2 mg/kg. Daily intraperitoneal administrations of 10 mg OA/kg resulted in 50% mortality by the tenth day of injection. Clinical symptoms included depression, huddling, roughened hair coats, humped backs and reduced weight gains. Mice injected intraperitoneally daily for 50 days with 5 mg OA/kg had a significantly (P<0.01) depressed antibody response to killed Brucella abortus. In contrast, oral administrations of OA at 4 ppm in feed for 50 days did not depress titre levels. Ochratoxin A also significantly (P<0.01 intraperitoneal; P<0.05 oral administrations) reduced body weight gain over the period of the trials. Neither oral nor intraperitoneal administration of OA for 50 days affected the response of mice to sheep red blood cells although both the number of antibody-forming cells and the number of cells per spleen were significantly lowered (P<0.01) by cyclophosphamide. Both spleen and body weights were significantly lowered (P<0.05) in the groups given OA. There was a significant depression of blast transformation (P<0.01) in mice treated intraperitoneally with either OA or cyclophosphamide and stimulated with concanavalin A; oral administration of OA did not depress blast transformation. It would appear that lower levels of exposure, e.g. 4 ppm OA in feed, do not cause depression of the immune response of mice. The depressive effect seen at much higher levels may be a result of a nonselective toxic effect. PMID:6804072

Mycotoxin production in wheat grains by different Aspergilli in relation to different relative humidities and storage periods.

PubMed

Atalla, Mohamed Mabrouk; Hassanein, Naziha Mohamed; El-Beih, Ahmed Atef; Youssef, Youssef Abdel-ghany

2003-02-01

Four different Aspergilli (Aspergillus oryzae, A. parasiticus, A. terreus and A. versicolor) were grown on wheat grains underdifferent degrees of relative humidity 14, 50, 74, 80 and 90%. Samples of wheat grains were taken monthly for a period of six months and examined for mycotoxin production. A. oryzae was found to produce aflatoxins B1, B2, zearalenone, DON and T-2 toxins under elevated degrees of humidity and prolonged periods of storage. A. parasiticus produced aflatoxins B1, G1, NIV, DON and T-2 toxins in high concentrations during a period of not more than three months storage at 14% relative humidity; at an increased level of relative humidity of 74% ochratoxin A, zearalenone and sterigmatocystin were also produced at high levels. The isolate was drastic in toxin production. A. terrus produced toxins at 14% relative humidity (aflatoxin G2 and DON) at levels much higher than any other prevalent degrees of humidity. A. versicolor is highly sensitive to relative humidity and grain moisture content It produced aflatoxins B1, G1, NIV and DON at a relative humidity of 50% and another toxins (aflatoxin G2, ochratoxins A, B and zearalenone) at 74%. The microorganism can be considered a trichothecene producer under suitable relative humidity.

Impact of pH on the stability and the cross-reactivity of ochratoxin A and citrinin.

PubMed

Bazin, Ingrid; Faucet-Marquis, Virginie; Monje, Marie-Carmen; El Khoury, Micheline; Marty, Jean-Louis; Pfohl-Leszkowicz, Annie

2013-11-28

Mycotoxins are secondary metabolites produced by several fungi contaminating crops. In several countries, the maximum permitted levels of mycotoxins are found in foodstuffs and feedstuffs. The common strategy of mycotoxin analysis involves extraction, clean-up and quantification by chromatography. In this paper, we analyzed the reasons of underestimation of ochratoxin A (OTA) content in wine, and overestimation of OTA in wheat, depending on the pH of the clean-up step and the simultaneous presence of citrinin (CIT). We demonstrated that the increase of pH by adding polyethylene glycol (PEG) to wine led to an underestimation of OTA by conversion of OTA into open ring ochratoxin A OP-OA. In comparing three methods of extraction and clean-up for the determination of OTA and CIT in wheat--(i) an inter-laboratory validated method for OTA in cereals using immunoaffinity column clean-up (IAC) and extraction by acetonitrile/water; (ii) a validated method using IAC and extraction with 1% bicarbonate Na; and (iii) an in-house validated method based on acid liquid/liquid extraction--we observed an overestimation of OTA after immunoaffinity clean-up when CIT is also present in the sample, whereas an underestimation was observed when OTA was alone. Under neutral and alkaline conditions, CIT was partially recognized by OTA antibodies.

Immunologic effects of low levels of ochratoxin A in ovo: utilization of a chicken embryo model.

PubMed

Harvey, R B; Kubena, L F; Naqi, S A; Gyimah, J E; Corrier, D E; Panigrahy, B; Phillips, T D

1987-01-01

Ochratoxin A (OA) was administered to 13-day-old chicken embryos via the chorioallantoic membrane. The 7-day LD50 value (day 20 incubation) of OA was calculated at 7.9 micrograms of OA. Ochratoxin-treated embryos (2.5 micrograms) had slight but significant changes in numbers of immunoglobulin-bearing cells in the bursa but not in the spleen. Chicks hatched from in ovo-treated eggs were challenged with 9 X 10(4) colony-forming units (CFU) of beta-hemolytic Escherichia coli (O1:K1) at 7 days of age via the thoracic air sac. Lesion scores of OA-treated chicks were equal to or less severe than those of controls. Hatchmates of the above chicks were vaccinated with a homologous killed E. coli bacterin (O1:K1) at both 2 and 4 weeks of age and challenged with 10(4) CFU of E. coli at 7 weeks. Post-challenge lesions were present in three vaccinated untreated controls and no OA-treated chicks. We conclude that although in ovo exposure to OA may marginally suppress immunoglobulin-bearing cells of bursa, chicks hatched from OA-treated eggs respond as well as controls to an antigen and resist infection by a virulent organism.

Impact of pH on the Stability and the Cross-Reactivity of Ochratoxin A and Citrinin

PubMed Central

Bazin, Ingrid; Faucet-Marquis, Virginie; Monje, Marie-Carmen; El Khoury, Micheline; Marty, Jean-Louis; Pfohl-Leszkowicz, Annie

2013-01-01

Mycotoxins are secondary metabolites produced by several fungi contaminating crops. In several countries, the maximum permitted levels of mycotoxins are found in foodstuffs and feedstuffs. The common strategy of mycotoxin analysis involves extraction, clean-up and quantification by chromatography. In this paper, we analyzed the reasons of underestimation of ochratoxin A (OTA) content in wine, and overestimation of OTA in wheat, depending on the pH of the clean-up step and the simultaneous presence of citrinin (CIT). We demonstrated that the increase of pH by adding polyethylene glycol (PEG) to wine led to an underestimation of OTA by conversion of OTA into open ring ochratoxin A OP-OA. In comparing three methods of extraction and clean-up for the determination of OTA and CIT in wheat—(i) an inter-laboratory validated method for OTA in cereals using immunoaffinity column clean-up (IAC) and extraction by acetonitrile/water; (ii) a validated method using IAC and extraction with 1% bicarbonate Na; and (iii) an in-house validated method based on acid liquid/liquid extraction—we observed an overestimation of OTA after immunoaffinity clean-up when CIT is also present in the sample, whereas an underestimation was observed when OTA was alone. Under neutral and alkaline conditions, CIT was partially recognized by OTA antibodies. PMID:24287570

Mycotoxins from mould infested building materials.

PubMed

Nielsen, K F

2000-03-01

Only limited documentation of non-allergenic, especially toxic reactions after inhalation of microfungal spores in water damaged buildings exists. Recently attention has been drawn to the mycotoxins as causal compounds, as some the dominating genera found in buildings are well known mycotoxin producers.Penicillium chrysogenum and A. ustus do not seem to produce any known mycotoxins when growing on building materials, whereasP. brevicompactum produces mycophenolic acid, someP. polonicum produces verrucosidin and verrucofortine,A. versicolor produces sterigmatocystins,A. niger produces nigragillin, orlandin, naphtho-γ-pyrones and tetracyclic compounds, someA. ochraceus produces ochratoxin A,Alternaria spp. produce alternariol and alternariol monomethyl ether,Chaetomium globosum produce chaetoglobosins, and finally 30-40% ofStachybotrys chartarum isolates from buildings produce macrocyclic trichothecenes and a number of other biologically active compounds.

Alkaloid cluster gene ccsA of the ergot fungus Claviceps purpurea encodes chanoclavine I synthase, a flavin adenine dinucleotide-containing oxidoreductase mediating the transformation of N-methyl-dimethylallyltryptophan to chanoclavine I.

PubMed

Lorenz, Nicole; Olsovská, Jana; Sulc, Miroslav; Tudzynski, Paul

2010-03-01

Ergot alkaloids are indole-derived secondary metabolites synthesized by the phytopathogenic ascomycete Claviceps purpurea. In wild-type strains, they are exclusively produced in the sclerotium, a hibernation structure; for biotechnological applications, submerse production strains have been generated by mutagenesis. It was shown previously that the enzymes specific for alkaloid biosynthesis are encoded by a gene cluster of 68.5 kb. This ergot alkaloid cluster consists of 14 genes coregulated and expressed under alkaloid-producing conditions. Although the role of some of the cluster genes in alkaloid biosynthesis could be confirmed by a targeted knockout approach, further functional analyses are needed, especially concerning the early pathway-specific steps up to the production of clavine alkaloids. Therefore, the gene ccsA, originally named easE and preliminarily annotated as coding for a flavin adenine dinucleotide-containing oxidoreductase, was deleted in the C. purpurea strain P1, which is able to synthesize ergot alkaloids in axenic culture. Five independent knockout mutants were analyzed with regard to alkaloid-producing capability. Thin-layer chromatography (TLC), ultrapressure liquid chromatography (UPLC), and mass spectrometry (MS) analyses revealed accumulation of N-methyl-dimethylallyltryptophan (Me-DMAT) and traces of dimethylallyltryptophan (DMAT), the first pathway-specific intermediate. Since other alkaloid intermediates could not be detected, we conclude that deletion of ccsA led to a block in alkaloid biosynthesis beyond Me-DMAT formation. Complementation with a ccsA/gfp fusion construct restored alkaloid biosynthesis. These data indicate that ccsA encodes the chanoclavine I synthase or a component thereof catalyzing the conversion of N-methyl-dimethylallyltryptophan to chanoclavine I.

[Toxicological characteristics of Ochratoxin A and its impact on male reproduction].

PubMed

Zhang, Tian-Yu; Zhao, Yong; Li, Lan; Shen, Wei

2017-08-01

Ochratoxin A (OTA) is found not only nephrotoxic, teratogenic, neurotoxic, and immunotoxic, but also reprotoxic for human and animals. In the recent decade, more attention has been paid to the impact of OTA on human reproduction and the studies of its underlying mechanisms. Many studies show that OTA affects the function of the reproductive system by acting as an endocrine disrupter and, as a testicular toxin, decreases sperm quality and even induces testis cancer. This review summarizes the toxicological characteristics and toxicokinetic process of OTA as well as recent progress in the studies of various toxic effects of OTA and their underlying mechanisms, hoping to call the attention from more people to the toxicity of OTA to male reproductive health.

Studies on carcinogenic and toxic effects of ochratoxin A in chicks.

PubMed

Stoev, Stoycho D

2010-04-01

Carcinogenic/toxic effects of ochratoxin A (OTA) in various internal organs of Plymouth Rock chicks were determined. The number of OTA-induced neoplasms was similar in chicks given 25 ppm L-β-phenylalanine (PHE) in addition to 5 ppm OTA compared to chicks given only 5 ppm OTA, which showed that PHE cannot be used as a real protector against the carcinogenic or toxic effects of OTA in chicks. OTA was found to provoke strong degenerative changes in liver and kidneys, degenerative changes and depletion of cells in lymphoid organs, oedematous and degenerative changes in the brain, muscular haemorrhages and fatty changes in the bone marrow. The target organs for carcinogenic effect of OTA in chicks were found to be kidneys and liver.

Ochratoxin A in Moroccan foods: occurrence and legislation.

PubMed

Zinedine, Abdellah

2010-05-01

Ochratoxin A (OTA) is secondary metabolite naturally produced in food and feed by toxigenic fungi, especially some Aspergillus species and Penicillium verucosum. OTA is one of the most studied mycotoxins and is of great interest due to its toxic effects on human and animals. OTA is produced in different food and feed matrices and contaminates a large range of base foods including cereals and derivatives, spices, dried fruits, wine and coffee, etc. Morocco, a North African country, has a climate characterized by high humidity and temperature, which probably favors the growth of molds. This contribution gives an overview of principal investigations about the presence of OTA in foods available in Morocco. Due to its toxicity, OTA presence is increasingly regulated worldwide, especially in countries of the European Union. However, up until now, no regulation limits were in force in Morocco, probably due to the ignorance of the health and economic problems resulting from OTA contamination. Finally, recommendations and future research directions are given required to assess the situation completely.

Ochratoxin A in Moroccan Foods: Occurrence and Legislation

PubMed Central

Zinedine, Abdellah

2010-01-01

Ochratoxin A (OTA) is secondary metabolite naturally produced in food and feed by toxigenic fungi, especially some Aspergillus species and Penicillium verucosum. OTA is one of the most studied mycotoxins and is of great interest due to its toxic effects on human and animals. OTA is produced in different food and feed matrices and contaminates a large range of base foods including cereals and derivatives, spices, dried fruits, wine and coffee, etc. Morocco, a North African country, has a climate characterized by high humidity and temperature, which probably favors the growth of molds. This contribution gives an overview of principal investigations about the presence of OTA in foods available in Morocco. Due to its toxicity, OTA presence is increasingly regulated worldwide, especially in countries of the European Union. However, up until now, no regulation limits were in force in Morocco, probably due to the ignorance of the health and economic problems resulting from OTA contamination. Finally, recommendations and future research directions are given required to assess the situation completely. PMID:22069630

Supercritical fluid extraction and direct fluid injection mass spectrometry for the determination of trichothecene mycotoxins in wheat samples

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kalinoski, H.T.; Udseth, H.R.; Wright, B.W.

1986-10-01

The application of on-line supercritical fluid extraction with chemical ionization mass spectrometry and collision induced dissociation tandem mass spectrometry for the rapid identification of parts-per-million levels of several trichothecene mycotoxins is demonstrated. Supercritical carbon dioxide is shown to allow identification of mycotoxins with minimum sample handling in complex natural matrices (e.g., wheat). Tandem mass spectrometry techniques are employed for unambiguous identification of compounds of varying polarity, and false positives from isobaric compounds are avoided. Capillary column supercritical fluid chromatography-mass spectrometry of a supercritical fluid extract of the same sample was also performed and detection limits in the parts-per-billion range appearmore » feasible.« less

Surveys of rice sold in Canada for aflatoxins, ochratoxin A and fumonisins

PubMed Central

Bansal, J.; Pantazopoulos, P.; Tam, J.; Cavlovic, P.; Kwong, K.; Turcotte, A.-M.; Lau, B.P.-Y.; Scott, P.M.

2011-01-01

Approximately 200 samples of rice (including white, brown, red, black, basmati and jasmine, as well as wild rice) from several different countries, including the United States, Canada, Pakistan, India and Thailand, were analysed for aflatoxins, ochratoxin A (OTA) and fumonisins by separate liquid Chromatographic methods in two different years. The mean concentrations for aflatoxin B1 (AFB1) were 0.19 and 0.17 ng g−1 with respective positive incidences of 56% and 43% (≥ the limit of detection (LOD) of 0.002 ng g−1). Twenty-three samples analysed in the second year also contained aflatoxin B2 (AFB2) at levels ≥LOD of 0.002 ng g−1 The five most contaminated samples in each year contained 1.44–7.14 ng AFB1 g−1 (year 1) and 1.45–3.48 ng AFB1 g−1 (year 2); they were mostly basmati rice from India and Pakistan and black and red rice from Thailand. The average concentrations of ochratoxin A (OTA) were 0.05 and 0.005 ng g−1 in year 1 and year 2, respectively; incidences of samples containing ≥LOD of 0.05 ng g−1 were 43% and 1%, respectively, in the 2 years. All positive OTA results were confirmed by LC-MS/MS. For fumonisins, concentrations of fumonisin B1 (FB1) averaged 4.5 ng g−1 in 15 positive samples (≥0.7 ng g−1) from year 1 (n = 99); fumonisin B2 (FB2) and fumonisin B3 (FB3) were also present (≥1 ng g−1). In the second year there was only one positive sample (14 ng g−1 FB1) out of 100 analysed. All positive FB1 results were confirmed by LC-MS/MS. PMID:21623501

[Biological contamination by micromycetes in dried Boletus edulis: research of aflatoxin B1, B2 G1, G2 and ochratoxin A].

PubMed

Lorini, C; Rossetti, F; Palazzoni, S; Comodo, N; Bonaccorsi, G

2008-01-01

Aim of this survey is to identify those filamentous fungi which parasite Boletus edulis and its group and check the potential presence of secondary metabolites, specifically aflatoxin B1, total aflatoxins and ochratoxin A, in order to assess the risk to consumers' health. Forty samples of dried Boletus edulis, collected by two food industries which distribute the product in many Italian regions, have been analysed. The sampling plan has been conducted from November 2005 to March 2006, collecting 50 g from each commercial category of dried Boletus edulis available in the factory at the time of sampling. All the samples have been tested by visual macroscopic and stereoscopic assays; for some samples--those referred to commercial category presumably at higher risk--we have performed cultural assays as well, typization of isolated micromycetes, extraction and quantification of aflatoxins and ochratoxin A. Mycotoxin detection has been made by HPLC, using the UNI EN 14123 and UNI EN 14132 standard methods, respectively applied to aflatoxins determination in peanuts, pistachios, figs and paprika and to ochratoxin A in barley and coffee. Non pathogenic micromycetes, common in food products, have been frequently observed in cultural assays, while Aspergillus flavus and Aspergillus niger have been found in some samples. However the concentration of aflatoxins was always under the quantification limit. The survey confirm that, if the cold chain is kept throughout the process and the distribution, Boletus edulis and analogue mycetes are not a favourable substratum for the growth and the development of moulds.

Identification and Characterization of Carboxylesterases from Brachypodium distachyon Deacetylating Trichothecene Mycotoxins

PubMed Central

Schmeitzl, Clemens; Varga, Elisabeth; Warth, Benedikt; Kugler, Karl G.; Malachová, Alexandra; Michlmayr, Herbert; Wiesenberger, Gerlinde; Mayer, Klaus F. X.; Mewes, Hans-Werner; Krska, Rudolf; Schuhmacher, Rainer; Berthiller, Franz; Adam, Gerhard

2015-01-01

Increasing frequencies of 3-acetyl-deoxynivalenol (3-ADON)-producing strains of Fusarium graminearum (3-ADON chemotype) have been reported in North America and Asia. 3-ADON is nearly nontoxic at the level of the ribosomal target and has to be deacetylated to cause inhibition of protein biosynthesis. Plant cells can efficiently remove the acetyl groups of 3-ADON, but the underlying genes are yet unknown. We therefore performed a study of the family of candidate carboxylesterases (CXE) genes of the monocot model plant Brachypodium distachyon. We report the identification and characterization of the first plant enzymes responsible for deacetylation of trichothecene toxins. The product of the BdCXE29 gene efficiently deacetylates T-2 toxin to HT-2 toxin, NX-2 to NX-3, both 3-ADON and 15-acetyl-deoxynivalenol (15-ADON) into deoxynivalenol and, to a lesser degree, also fusarenon X into nivalenol. The BdCXE52 esterase showed lower activity than BdCXE29 when expressed in yeast and accepts 3-ADON, NX-2, 15-ADON and, to a limited extent, fusarenon X as substrates. Expression of these Brachypodium genes in yeast increases the toxicity of 3-ADON, suggesting that highly similar genes existing in crop plants may act as susceptibility factors in Fusarium head blight disease. PMID:26712789

Alkaloid Cluster Gene ccsA of the Ergot Fungus Claviceps purpurea Encodes Chanoclavine I Synthase, a Flavin Adenine Dinucleotide-Containing Oxidoreductase Mediating the Transformation of N-Methyl-Dimethylallyltryptophan to Chanoclavine I ▿

PubMed Central

Lorenz, Nicole; Olšovská, Jana; Šulc, Miroslav; Tudzynski, Paul

2010-01-01

Ergot alkaloids are indole-derived secondary metabolites synthesized by the phytopathogenic ascomycete Claviceps purpurea. In wild-type strains, they are exclusively produced in the sclerotium, a hibernation structure; for biotechnological applications, submerse production strains have been generated by mutagenesis. It was shown previously that the enzymes specific for alkaloid biosynthesis are encoded by a gene cluster of 68.5 kb. This ergot alkaloid cluster consists of 14 genes coregulated and expressed under alkaloid-producing conditions. Although the role of some of the cluster genes in alkaloid biosynthesis could be confirmed by a targeted knockout approach, further functional analyses are needed, especially concerning the early pathway-specific steps up to the production of clavine alkaloids. Therefore, the gene ccsA, originally named easE and preliminarily annotated as coding for a flavin adenine dinucleotide-containing oxidoreductase, was deleted in the C. purpurea strain P1, which is able to synthesize ergot alkaloids in axenic culture. Five independent knockout mutants were analyzed with regard to alkaloid-producing capability. Thin-layer chromatography (TLC), ultrapressure liquid chromatography (UPLC), and mass spectrometry (MS) analyses revealed accumulation of N-methyl-dimethylallyltryptophan (Me-DMAT) and traces of dimethylallyltryptophan (DMAT), the first pathway-specific intermediate. Since other alkaloid intermediates could not be detected, we conclude that deletion of ccsA led to a block in alkaloid biosynthesis beyond Me-DMAT formation. Complementation with a ccsA/gfp fusion construct restored alkaloid biosynthesis. These data indicate that ccsA encodes the chanoclavine I synthase or a component thereof catalyzing the conversion of N-methyl-dimethylallyltryptophan to chanoclavine I. PMID:20118373

Combinatorial assembly of simple and complex D-lysergic acid alkaloid peptide classes in the ergot fungus Claviceps purpurea.

PubMed

Ortel, Ingo; Keller, Ullrich

2009-03-13

The ergot fungus Claviceps purpurea produces both ergopeptines and simple d-lysergic acid alkylamides. In the ergopeptines, such as ergotamine, d-lysergic acid is linked to a bicyclic tripeptide in amide-like fashion, whereas in the d-lysergylalkanolamides it is linked to an amino alcohol derived from alanine. We show here that these compound classes are synthesized by a set of three non-ribosomal lysergyl peptide synthetases (LPSs), which interact in a combinatorial fashion for synthesis of the relevant product. The trimodular LPS1 assembles with LPS2, the d-lysergic acid recruiting module, to synthesize the d-lysergyltripeptide precursors of ergopeptines from d-lysergic acid and the three amino acids of the peptide chain. Alternatively, LPS2 can assemble with a distinct monomodular non-ribosomal peptide synthetase (NRPS) subunit (ergometrine synthetase) to synthesize the d-lysergic acid alkanolamide ergometrine from d-lysergic acid and alanine. The synthesis proceeds via covalently bound d-lysergyl alanine and release of dipeptide as alcohol with consumption of NADPH. Enzymatic and immunochemical analyses showed that ergometrine synthetase is most probably the enzyme LPS3 whose gene had been identified previously as part of the ergot alkaloid biosynthesis gene cluster in C. purpurea. Inspections of all LPS sequences showed no recognizable peptide linkers for their protein-protein interactions as in NRPS subunits of bacteria. Instead, they all carry conserved N-terminal domains (C0-domains) with similarity to the C-terminal halves of NRPS condensation domains pointing to an alternative mechanism of subunit-subunit interactions in fungal NRPS systems. Phylogenetic analysis of LPS modules and the C0-domains suggests that these enzyme systems most probably evolved by module duplications and rearrangements from a bimodular ancestor.

Label-Free G-Quadruplex Aptamer Fluorescence Assay for Ochratoxin A Using a Thioflavin T Probe.

PubMed

Wu, Kefeng; Ma, Changbei; Zhao, Han; He, Hailun; Chen, Hanchun

2018-05-12

Ochratoxin A (OTA) is one of the most common mycotoxins contaminating feed and foodstuffs. Therefore, a great deal of concern is associated with AFB1 toxicity. In this work, a fast and sensitive fluorescence aptamer biosensor has been proposed for the OTA assay. In the absence of OTA, the OTA aptamer can form a G-quadruplex structure with thioflavin T (ThT) dye, which results in increased fluorescence. After joining OTA, OTA aptamer combines with OTA and the G-quadruplex can be formed. Only faint fluorescence was finally observed when ThT weakly reacts with the quadruplex. Through this test method, the entire reaction and analysis process of OTA can be completed in 10 min. Under optimal experimental conditions (600 nM OTA-APT, 7 μM ThT, and 3 min incubation time), this proposed assay has a good limit of detection (LOD) of 0.4 ng/mL and shows a good linear relationship within the range of 1.2⁻200 ng/mL under the best experimental conditions. This method has a high specificity for OTA relative to Ochratoxin B (23%) and Aflatoxin B₁ (13%). In addition, the quantitative determination of this method in real samples has been validated using a sample of red wine supplemented with a range of OTA concentrations (1.2 ng/mL, 12 ng/mL, and 40 ng/mL) with recoveries of 96.5% to 107%.

Dietary exposure to aflatoxin B1, ochratoxin A and fuminisins of adults in Lao Cai province, Viet Nam: A total dietary study approach.

PubMed

Huong, Bui Thi Mai; Tuyen, Le Danh; Tuan, Do Huu; Brimer, Leon; Dalsgaard, Anders

2016-12-01

Aflatoxins, fumonisins and ochratoxin A that contaminate various agricultural commodities are considered of significant toxicity and potent human carcinogens. This study took a total dietary study approach and estimated the dietary exposure of these mycotoxins for adults living in Lao Cai province, Vietnam. A total of 42 composite food samples representing 1134 individual food samples were prepared according to normal household practices and analysed for the three mycotoxins. Results showed that the dietary exposure to aflatoxin B1 (39.4 ng/kg bw/day) and ochratoxin A (18.7 ng/kg bw/day) were much higher than recommended provisional tolerable daily intake (PTDI) values mainly due to contaminated cereals and meat. The exposure to total fumonisins (1400 ng/kg bw/day) was typically lower than the PTDI value (2000 ng/kg bw/day). The estimated risk of liver cancer associated with exposure to aflatoxin B1 was 2.7 cases/100,000 person/year. Margin of exposure (MOE) of renal cancer linked to ochratoxin A and liver cancer associated with fumonisins were 1124 and 1954, respectively indicating risk levels of public health concern. Further studies are needed to evaluate the efficiency of technical solutions which could reduce mycotoxin contamination as well as to determine the health effects of the co-exposure to different types of mycotoxins. Copyright © 2016 Elsevier Ltd. All rights reserved.

Occurrence of ochratoxin A in wine and beer samples from China.

PubMed

Wu, Jianwei; Tan, Yinfeng; Wang, Yuqi; Xu, Rong

2011-01-01

A total of 125 wine and beer samples, including 42 white, 63 red wine and 20 beer samples, originating from several areas of China were analyzed for ochratoxin A (OTA). An analytical method based on immunoaffinity column (IAC) for clean-up and high performance liquid chromatography with fluorescence detection (HPLC-FD) was used to determine OTA. Positive results were further confirmed by LC-MS-MS. OTA was detected in 22 (17.6%) samples at levels of 0.02-1.18 ng/ml, which were below the EU maximum limit. The mean OTA concentration in red wine was slightly higher than in white wine. This study showed that OTA occurs in wine and beer in China but at levels that pose no danger of significant human exposure to OTA from this source.

The bZIP Transcription Factor Fgap1 Mediates Oxidative Stress Response and Trichothecene Biosynthesis But Not Virulence in Fusarium graminearum

PubMed Central

Montibus, Mathilde; Ducos, Christine; Bonnin-Verdal, Marie-Noelle; Bormann, Jorg; Ponts, Nadia; Richard-Forget, Florence; Barreau, Christian

2013-01-01

Redox sensing is of primary importance for fungi to cope with oxidant compounds found in their environment. Plant pathogens are particularly subject to the oxidative burst during the primary steps of infection. In the budding yeast Saccharomyces cerevisiae, it is the transcription factor Yap1 that mediates the response to oxidative stress via activation of genes coding for detoxification enzymes. In the cereal pathogen Fusarium graminearum, Fgap1 a homologue of Yap1 was identified and its role was investigated. During infection, this pathogen produces mycotoxins belonging to the trichothecenes family that accumulate in the grains. The global regulation of toxin biosynthesis is not completely understood. However, it is now clearly established that an oxidative stress activates the production of toxins by F. graminearum. The involvement of Fgap1 in this activation was investigated. A deleted mutant and a strain expressing a truncated constitutive form of Fgap1 were constructed. None of the mutants was affected in pathogenicity. The deleted mutant showed higher level of trichothecenes production associated with overexpression of Tri genes. Moreover activation of toxin accumulation in response to oxidative stress was no longer observed. Regarding the mutant with the truncated constitutive form of Fgap1, toxin production was strongly reduced. Expression of oxidative stress response genes was not activated in the deleted mutant and expression of the gene encoding the mitochondrial superoxide dismutase MnSOD1 was up-regulated in the mutant with the truncated constitutive form of Fgap1. Our results demonstrate that Fgap1 plays a key role in the link between oxidative stress response and F. graminearum secondary metabolism. PMID:24349499

Brachypodium distachyon as alternative model host system for the ergot fungus Claviceps purpurea.

PubMed

Kind, Sabine; Schurack, Selma; Hinsch, Janine; Tudzynski, Paul

2018-04-01

To investigate its susceptibility to ergot infection, we inoculated Brachypodium distachyon with Claviceps purpurea and compared the infection symptoms with those on rye (Secale cereale). We showed that, after inoculation of Brachypodium with Claviceps, the same disease symptoms occurred in comparable temporal and spatial patterns to those on rye. The infection rate of Claviceps on this host was reduced compared with rye, but the disease could be surveyed by fungal genomic DNA quantification. Mutants of Claviceps which were virulence attenuated on rye were also affected on Brachypodium. We were able to show that pathogenesis-related gene expression changed in a typical manner for biotrophic pathogen attack. Our results indicated that the Claviceps-Brachypodium interaction was dependent on salicylic acid, cytokinin and auxin. We consider Brachypodium to be a suitable and useful alternative host; the increased sensitivity compared with rye will be valuable for the identification of infection mechanisms. Future progess in understanding the Claviceps-plant interaction will be facilitated by the use of a well-characterized model host system. © 2017 BSPP AND JOHN WILEY & SONS LTD.

Pharmacotherapy for organic brain syndrome in late life. Evaluation of an ergot derivative vs placebo.

PubMed

Gaitz, C M; Varner, R V; Overall, J E

1977-07-01

Evaluation of treatment modalities, including pharmacotherapy, for organic brain syndrome (OBS) has been difficult because of sampling and methodological problems, and comparisons of research studies are all but impossible. In this study, an ergot derivative, a combination of dihydroergocornine mesylate, dihydroergocristine mesylate, and dihydroergokryptine mesylate (Hydergine) was compared with placebo, using a double-blind technique in a sample of nursing home residents with evidence of OBS. An 18-category symptom rating scale was used for periodic assessment over a six-month interval. Comparisons of the two groups of subjects disclosed that the Hydergine-treated group showed statistically significantly more improvement in most of the variables measured, especially during the last three months of treatment. Furthermore, sophisticated analysis revealed that positive changes in cognitive function cannot be accounted for as a mere reflection, or "halo" effect, associated with improved mood and general sense of well-being.

Molecular cloning and analysis of the ergopeptine assembly system in the ergot fungus Claviceps purpurea.

PubMed

Correia, Telmo; Grammel, Nicolas; Ortel, Ingo; Keller, Ullrich; Tudzynski, Paul

2003-12-01

Claviceps purpurea produces the pharmacological important ergopeptines, a class of cyclol-structured alkaloid peptides containing D-lysergic acid. These compounds are assembled from D-lysergic acid and three different amino acids by the nonribosomal peptide synthetase enzymes LPS1 and LPS2. Cloning of alkaloid biosynthesis genes from C. purpurea has revealed a gene cluster including two NRPS genes, cpps 1 and cpps 2. Protein sequence data had assigned earlier cpps1 to encode the trimodular LPS1 assembling the tripeptide portion of ergopeptines. Here, we show by transcriptional analysis, targeted inactivation, analysis of disruption mutants, and heterologous expression that cpps 2 encodes the monomodular LPS2 responsible for D-lysergic acid activation and incorporation into the ergopeptine backbone. The presence of two distinct NRPS subunits catalyzing formation of ergot peptides is the first example of a fungal NRPS system consisting of different NRPS subunits.

An overview of the toxicology and toxicokinetics of fusarenon-X, a type B trichothecene mycotoxin.

PubMed

Aupanun, Sawinee; Poapolathep, Saranya; Giorgi, Mario; Imsilp, Kanjana; Poapolathep, Amnart

2017-01-20

Fusarenon-X (FX) is a type B trichothecene mycotoxin that is frequently observed along with deoxynivalenol (DON) and nivalenol (NIV) in agricultural commodities. This review aims to give an overview of the literature concerning the toxicology and toxicokinetics of FX. FX is primarily found in cereals grown in temperate regions, but it can also be found worldwide because of the global transport of products. The major toxicity of FX occurs through inhibition of protein synthesis, followed by the disruption of DNA synthesis. Moreover, FX has also been shown to induce apoptosis in in vitro and in vivo studies. The targets of FX are organs containing actively proliferating cells, such as the thymus, spleen, skin, small intestine, testes and bone marrow. FX causes immunosuppression, intestinal malabsorption, developmental toxicity and genotoxicity. In addition, sufficient evidence of carcinogenicity in experimental animals is currently lacking, and the International Agency for Research on Cancer (IARC) classifies it as a group 3 carcinogen.

Mycobiota and Natural Incidence of Aflatoxins, Ochratoxin A, and Citrinin in Indian Spices Confirmed by LC-MS/MS

PubMed Central

Jeswal, Punam; Kumar, Dhiraj

2015-01-01

Nine different Indian spices (red chilli, black pepper, turmeric, coriander, cumin, fennel, caraway, fenugreek, and dry ginger) commonly cultivated and highly used in India were analysed for natural occurrence of toxigenic mycoflora and aflatoxins (AFs), ochratoxin A (OTA), and citrinin (CTN) contamination. Aspergillus flavus and Aspergillus niger were the most dominant species isolated from all types of spices. Red chilli samples were highly contaminated with aflatoxins (85.4%) followed by dry ginger (77.7%). 56% Aspergillus flavus from red chilli and 45% Aspergillus ochraceus from black pepper were toxigenic and produced aflatoxins and ochratoxin A, respectively. Qualitative detection and quantitative detection of mycotoxins in spices were analyzed by ELISA and further confirmed by LC-MS/MS. Penicillium citrinum produced citrinin in red chilli, black pepper, coriander, cumin, fenugreek, and dry ginger samples. The highest amount of AFs was found in red chilli (219.6 ng/g), OTA was in black pepper (154.1 ng/g), and CTN was in dry ginger samples (85.1 ng/g). The results of this study suggest that the spices are susceptible substrate for growth of mycotoxigenic fungi and further mycotoxin production. This is the first report of natural occurrence of citrinin in black pepper and dry ginger from India. PMID:26229535

Colorimetric Analysis of Ochratoxin A in Beverage Samples

PubMed Central

Bueno, Diana; Valdez, Luis F.; Gutiérrez Salgado, Juan Manuel; Marty, Jean Louis; Muñoz, Roberto

2016-01-01

This manuscript describes the use of a portable and low cost fluorescence setup to quantify the concentration of ochratoxin A (OTA) in beverage samples using an in-house developed system and different color models. It is reported that OTA is naturally fluorescent, for that reason an ultraviolet light at 365 nm was used to excite the samples and a Complementary Metal Oxide Semiconductor (CMOS) sensor was used to get a photograph of the OTA under excitation conditions, which is controlled by an executable interface designed in MATLAB. For each concentration of OTA, the coordinates with respect to each model color were obtained and plotted to quantify the mycotoxin present in the sample. It was possible to observe that despite the fact no extraction column was employed, the Red, Green, Blue (RGB) model shows a proportional relation to the evaluated concentrations. Despite the fact more analysis and other methods are required to quantify the OTA concentration, the brightness and a,b for the color-opponent dimensions (L*a*b) and Hue, Saturation, Value (HSV) tests provide results whereby it is possible to identify the concentration of OTA in beverage samples such as beer and wine. PMID:27834900

Combinatorial Assembly of Simple and Complex d-Lysergic Acid Alkaloid Peptide Classes in the Ergot Fungus Claviceps purpurea*S⃞

PubMed Central

Ortel, Ingo; Keller, Ullrich

2009-01-01

The ergot fungus Claviceps purpurea produces both ergopeptines and simple d-lysergic acid alkylamides. In the ergopeptines, such as ergotamine, d-lysergic acid is linked to a bicyclic tripeptide in amide-like fashion, whereas in the d-lysergylalkanolamides it is linked to an amino alcohol derived from alanine. We show here that these compound classes are synthesized by a set of three non-ribosomal lysergyl peptide synthetases (LPSs), which interact in a combinatorial fashion for synthesis of the relevant product. The trimodular LPS1 assembles with LPS2, the d-lysergic acid recruiting module, to synthesize the d-lysergyltripeptide precursors of ergopeptines from d-lysergic acid and the three amino acids of the peptide chain. Alternatively, LPS2 can assemble with a distinct monomodular non-ribosomal peptide synthetase (NRPS) subunit (ergometrine synthetase) to synthesize the d-lysergic acid alkanolamide ergometrine from d-lysergic acid and alanine. The synthesis proceeds via covalently bound d-lysergyl alanine and release of dipeptide as alcohol with consumption of NADPH. Enzymatic and immunochemical analyses showed that ergometrine synthetase is most probably the enzyme LPS3 whose gene had been identified previously as part of the ergot alkaloid biosynthesis gene cluster in C. purpurea. Inspections of all LPS sequences showed no recognizable peptide linkers for their protein-protein interactions as in NRPS subunits of bacteria. Instead, they all carry conserved N-terminal domains (C0-domains) with similarity to the C-terminal halves of NRPS condensation domains pointing to an alternative mechanism of subunit-subunit interactions in fungal NRPS systems. Phylogenetic analysis of LPS modules and the C0-domains suggests that these enzyme systems most probably evolved by module duplications and rearrangements from a bimodular ancestor. PMID:19139103

Early harvest and ensilage of forage sorghum infected with ergot (Claviceps africana) reduces the risk of livestock poisoning.

PubMed

Blaney, B J; Ryley, M J; Boucher, B D

2010-08-01

Sorghum ergot produces dihydroergosine (DHES) and related alkaloids, which cause hyperthermia in cattle. Proportions of infected panicles (grain heads), leaves and stems were determined in two forage sorghum crops extensively infected 2 to 4 weeks prior to sampling and the panicles were assayed for DHES. Composite samples from each crop, plus a third grain variety crop, were coarsely chopped and half of each sealed in plastic buckets for 6 weeks to simulate ensilation. The worst-infected panicles contained up to 55 mg DHES/kg, but dilution reduced average concentrations of DHES in crops to approximately 1 mg/kg, a relatively safe level for cattle. Ensilation significantly (P = 0.043) reduced mean DHES concentrations from 0.85 to 0.46 mg/kg.

Label-Free QCM Immunosensor for the Detection of Ochratoxin A

PubMed Central

Ertekin, Özlem; Laguna, Duygu Ercan; Özen, Fehime Şeyma; Öztürk, Zafer Ziya; Öztürk, Selma

2018-01-01

Ochratoxin A (OTA) is a potent mycotoxin that poses a risk in food and feed moieties and subject to worldwide regulation. Laboratory-based analytical methods are traditionally employed for reliable OTA quantification, but these methods cannot provide rapid and on-site analysis, where biosensors fill this gap. In this study a label-free quartz crystal microbalance (QCM)-based immunosensor for the detection of OTA, which is one of the most important small molecule contaminants, was developed by direct immobilization of OTA to amine-bearing sensor surfaces using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC)/N-Hydroxysuccinimide (NHS) chemistry. The protein-free sensor surface enabled regeneration of sensor surface with 50 mM NaOH and 1% SDS up to 13 times without loss of performance, which would disrupt a protein-containing sensor surface. We developed a QCM immunosensor using the developed sensor surface with a 17.2–200 ng/mL detection range which can be used for on-site detection of feedstuffs. PMID:29641432

Label-Free QCM Immunosensor for the Detection of Ochratoxin A.

PubMed

Pirinçci, Şerife Şeyda; Ertekin, Özlem; Laguna, Duygu Ercan; Özen, Fehime Şeyma; Öztürk, Zafer Ziya; Öztürk, Selma

2018-04-11

Ochratoxin A (OTA) is a potent mycotoxin that poses a risk in food and feed moieties and subject to worldwide regulation. Laboratory-based analytical methods are traditionally employed for reliable OTA quantification, but these methods cannot provide rapid and on-site analysis, where biosensors fill this gap. In this study a label-free quartz crystal microbalance (QCM)-based immunosensor for the detection of OTA, which is one of the most important small molecule contaminants, was developed by direct immobilization of OTA to amine-bearing sensor surfaces using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC)/N-Hydroxysuccinimide (NHS) chemistry. The protein-free sensor surface enabled regeneration of sensor surface with 50 mM NaOH and 1% SDS up to 13 times without loss of performance, which would disrupt a protein-containing sensor surface. We developed a QCM immunosensor using the developed sensor surface with a 17.2-200 ng/mL detection range which can be used for on-site detection of feedstuffs.

A Label-Free Aptasensor for Ochratoxin a Detection Based on the Structure Switch of Aptamer.

PubMed

Liu, Feng; Ding, Ailing; Zheng, Jiushang; Chen, Jiucun; Wang, Bin

2018-06-01

A label-free sensing platform is developed based on switching the structure of aptamer for highly sensitive and selective fluorescence detection of ochratoxin A (OTA). OTA induces the structure of aptamer, transforms into G-quadruplex and produces strong fluorescence in the presence of zinc(II)-protoporphyrin IX probe due to the specific bind to G-quadruplex. The simple method exhibits high sensitivity towards OTA with a detection limit of 0.03 nM and excellent selectivity over other mycotoxins. In addition, the successful detection of OTA in real samples represents a promising application in food safety.

Influence of tryptophan and related compounds on ergot alkaloid formation in Claviceps purpurea (FR.) Tul.

PubMed

Erge, D; Schumann, B; Gröger, D

1984-01-01

L-Tryptophan did not exert any influence on peptide alkaloid formation in an ergotamine and in an ergosine-accumulating C. purpurea strain. A different picture was observed in a series of related C. purpurea strains. Tryptophan showed a slight stimulatory effect on the ergotoxine producer Pepty 695/S. A blocked mutant of it, designated as Pepty 695/ch which was able to accumulate secoclavines gave similar results. In a high-yielding elymoclavine strain Pepty 695/e, the progeny of the former one, tryptophan up to a concentration of 25 mM stimulated remarkably clavine biosynthesis. Furthermore, tryptophan could overcome the block of synthesis by inorganic phosphate. Increased specific activities of chanoclavine cyclase but not DMAT synthetase were observed in cultures of strain Pepty 695/e supplemented with tryptophan. 5-Methyltryptophan and bioisosteres of tryptophan were ineffective in alkaloid stimulation. These results are compared with those obtained with the grass ergot strain SD 58 and discussed with the relation to other induction phenomena.

Ochratoxin A levels in spices and dried nuts consumed in Tunisia.

PubMed

Zaied, Chiraz; Abid, Salwa; Bouaziz, Chayma; Chouchane, Salwa; Jomaa, Mohamed; Bacha, Hassen

2010-01-01

A total of 112 samples of spices (24 caraway, 20 coriander, 25 curcuma, 20 black pepper and 23 red pepper) and 110 samples of dried nuts (44 almonds, 42 peanuts and 24 pistachio) purchased from popular markets in 24 regions of Tunisia were analyzed for ochratoxin A (OTA) by fluorescence HPLC. The average levels of contamination of OTA found in spice samples were 244, 206, 290, 274 and 203 µg/kg, respectively, for caraway, coriander, curcuma, black pepper and red pepper. Concerning dried nut samples, the average levels were 61, 60 and 89 µg/kg, respectively, for almonds, peanuts and pistachio. Contamination levels were higher than the usual norms (10.0 OTA µg/kg) established by the European Commission in 2005 . This survey is the first to be carried out on the natural occurrence of OTA in the main spices and dried nuts consumed by the Tunisian population.

Degradation and epimerization of ergot alkaloids after baking and in vitro digestion.

PubMed

Merkel, Stefan; Dib, Baha; Maul, Ronald; Köppen, Robert; Koch, Matthias; Nehls, Irene

2012-11-01

The degradation and epimerization of ergot alkaloids (EAs) in rye flour were investigated after baking cookies and subsequently subjecting them to an in vitro digestion model. Different steps of digestion were analyzed using salivary, gastric, and duodenal juices. The degradation and bidirectional conversion of the toxicologically relevant (R)-epimers and the biologically inactive (S)-epimers for seven pairs of EAs were determined by a HPLC method coupled with fluorescence detection. Baking cookies resulted in degradation of EAs (2-30 %) and a shift in the epimeric ratio toward the (S)-epimer for all EAs. The applied digestion model led to a selective toxification of ergotamine and ergosine, two ergotamine-type EAs. The initial percentage of the toxic (R)-epimer in relation to the total toxin content was considerably increased after digestion of cookies. Ergotamine and ergosine increased from 32 to 51 % and 35 to 55 %, respectively. In contrast, EAs of the ergotoxine type (ergocornine, α- and β-ergocryptine, and ergocristine) showed an epimeric shift toward their biologically inactive (S)-epimers. Further experiments indicated that the selective epimerization of ergotamine EAs occurs in the duodenal juice only. These results demonstrate that toxification of EAs in the intestinal tract should be taken into consideration.

Efficacy of different caffeine concentrations on growth and ochratoxin A production by Aspergillus species.

PubMed

Akbar, A; Medina, A; Magan, N

2016-07-01

The objective of this study was to evaluate the effect of different caffeine concentrations (0-4%) on (i) lag phase prior to growth, (ii) growth rates and (iii) ochratoxin A (OTA) production by strains from the Aspergillus section Circumdati and Aspergillus section Nigri groups, isolated from coffee, when grown on a conducive medium at 0·98 water activity and 30°C. The lag phases prior to growth increased with caffeine concentration. A strain of Aspergillus niger and Aspergillus carbonarius were the most sensitive to caffeine with growth being inhibited by <1% caffeine. For strains of Aspergillus westerdijkiae, Aspergillus ochraceus and Aspergillus steynii, although growth was inhibited significantly, some growth (10-15% of controls) occurred in 4% caffeine. OTA production was significantly inhibited by only 0·5% caffeine for strains of A. westerdijkiae, A. niger and A. carbonarius. For A. steynii at least 1·5% caffeine was required to inhibit OTA production. In contrast, for the strain of A. ochraceus there was a stimulation of OTA at 3% with a reduction at 4% caffeine. These results are discussed in the context of the different concentrations of caffeine found in Arabica and Robusta coffee and the development of minimization strategies. Arabic (0·6%) and Robusta coffee (4%) have significantly different amounts of endogenous caffeine. The growth of six ochratoxigenic fungi which contaminate coffee with ochratoxin A (OTA) had differential tolerance/sensitivity to concentrations of caffeine in vitro in this range. However, low concentrations of caffeine (<0·5%) was inhibitory to OTA production. These results are discussed in the context of the potential for using such information for the design of minimization strategies to control mycotoxin production in such products. © 2016 The Society for Applied Microbiology.

An electrochemical immunosensor for ochratoxin A determination in wines based on a monoclonal antibody and paramagnetic microbeads.

PubMed

Vidal, Juan C; Bonel, Laura; Ezquerra, Alba; Duato, Patricia; Castillo, Juan R

2012-06-01

We report a direct competitive immunosensor for the rapid determination of ochratoxin A (OTA) in wine samples. Magnetic beads (1 ± 0.5 μm diameter) covered with streptavidin were functionalized with a monoclonal antibody against OTA, and then left to incubate in a solution of tracer (ochratoxin conjugated to the enzyme peroxidase) and a range of OTA concentrations (10(-4) to 1,000 ng mL(-1)). After washing and separation steps helped with a magnetic field, a volume of the dispersion was put on screen-printed electrodes under a magnet, and after adding the substrate the p-benzoquinone generated enzymatically was detected by differential-pulse voltammetry. Wine samples (2 mL) were easily prepared simply by adjusting to pH = 7.5 with diluted NaOH and by adding polyvinylpyrrolidone for complexing polyphenols, without any other clean-up or preconcentration steps. The limit of detection for detecting OTA in wines was of 0.11 ± 0.01 ng L(-1), well below the permitted content of the mycotoxin by the European Union (<2 ng mL(-1)). Spiked wines were subjected to immunosensor calibrations to study the matrix effects. OTA concentrations measured with the immunosensor were compared with those obtained by high-performance liquid chromatography coupled to fluorescence detection (AOAC official method 2001.01). The OTA levels from two red wines of "Campo de Borja", Spain, ranged from about 0.027 to 0.033 ng mL(-1) of OTA.

An optical fiber-based LSPR aptasensor for simple and rapid in-situ detection of ochratoxin A.

PubMed

Lee, Bobin; Park, Jin-Ho; Byun, Ju-Young; Kim, Joon Heon; Kim, Min-Gon

2018-04-15

Label-free biosensing methods that rely on the use of localized surface plasmon resonance (LSPR) have attracted great attention as a result of their simplicity, high sensitivity, and relatively low cost. However, in-situ analysis of real samples using these techniques has remained challenging because colloidal nanoparticles (NPs) can be unstable at certain levels of pH and salt concentration. Even in the case of a chip-type LSPR sensor that can resolve the instability problem by employing NPs immobilized on the substrate, loading of a sample to sensor chip with exact volume control can be difficult for unskilled users. Herein, we report an optical-fiber-based LSPR aptasensor that can avoid these problems and serve as a portable and simple system for sensitive detection of a small mycotoxin, ochratoxin A (OTA), in real samples. The optical fiber coated with aptamer-modified gold nanorods (GNRs) is simply dipped into a solution containing OTA and subjected to LSPR analysis. Quantitative analysis of OTA is performed by measuring the spectral red shift of the LSPR peak of GNRs. Under optimized conditions, the LSPR peak shift displays a linear response (R 2 = 0.9887) to OTA in the concentration range from 10pM to 100nM, with a limit of detection of 12.0pM (3S). The developed sensor shows a high selectivity for OTA over other mycotoxins such as zearalenone (ZEN) and ochratoxin B (OTB), and shows an accurate detection capability for OTA in real grape juice samples. Copyright © 2017 Elsevier B.V. All rights reserved.

Survey of Slaughtered Pigs for Occurrence of Ochratoxin A and Porcine Nephropathy in Serbia

PubMed Central

Milićević, Dragan; Jurić, Verica; Stefanović, Srđan; Jovanović, Milijan; Janković, Saša

2008-01-01

Samples of blood, kidney and liver were randomly selected from slaughtered pigs (n=90) and analyzed for ochratoxin A by HPLC. In addition, in order to obtain information on the occurrence of nephropathy, histological examinations were carried out. Of the 90 liver samples, 26.6% contained OTA in the range of 0.22–14.5 ng/g. The incidence of OTA in serum and kidney were very similar (31%, 33.3%), with a maximum concentration of 220.8 ng/mL, and 52.5 ng/g, respectively. Histopathological examination of kidneys confirmed tubulopathies with edema and cell vacuolization. In addition, hemorrhages and necrosis of proximal kidney tubules’ cells were found. PMID:19330066

Inhibition of protein synthesis in maize and wheat by trichothecene mycotoxins and hybridoma-based enzyme immunoassay for deoxynivalenol

DOE Office of Scientific and Technical Information (OSTI.GOV)

Casale, W.L.

1987-01-01

The 12,13-epoxytrichothecene mycotoxins deoxynivalenol (DON, vomitoxin) and T-2 toxin inhibited protein synthesis in vivo and in cell-free systems from wheat and maize, host plants of trichothecene-producing Fusarium spp.Protein synthesis in tissue (leaf discs and kernel sections) was measured by incorporation of /sup 3/H-leucine into acetone:ethanol insoluble material, and in cell-free translation systems from wheat embryos and maize seedling plumules by incorporation of /sup 3/H-leucine into trichloroacetic acid-insoluble material. The toxin concentration inhibiting 50% of /sup 3/H-leucine incorporation (ID/sub 50/) by several maize varieties were 0.9 ..mu..M (T-2 toxin) and 9-22 ..mu..M (DON). ID/sub 50/ values for wheat were 0.25 ..mu..Mmore » (T-2 toxin) and 4.5 ..mu..M (DON).« less

Electrochemical Aptamer-Based Sensors for Rapid Point-of-Use Monitoring of the Mycotoxin Ochratoxin A Directly in a Food Stream.

PubMed

Somerson, Jacob; Plaxco, Kevin W

2018-04-15

The ability to measure the concentration of specific small molecules continuously and in real-time in complex sample streams would impact many areas of agriculture, food safety, and food production. Monitoring for mycotoxin taint in real time during food processing, for example, could improve public health. Towards this end, we describe here an inexpensive electrochemical DNA-based sensor that supports real-time monitor of the mycotoxin ochratoxin A in a flowing stream of foodstuffs.

Effect of Robusta (Coffea canephora P.) coffee cherries quantity put out for sun drying on contamination by fungi and ochratoxin A (OTA) under tropical humid zone (Côte d'Ivoire).

PubMed

Kouadio, Irène Ahou; Koffi, Louis Ban; Nemlin, Jean Gnopo; Dosso, Mireille Bretin

2012-06-01

The effect of coffee cherries quantity put out for sun drying on the kinetics of the drying, chemical components variation, fungal growth and ochratoxin A production was evaluated. The results showed that the more coffee cherries quantity on the drying area was important, the slower they dried. Indeed, the drying durations were 12, 17, 21, 26, 31 and 32 days respectively for the lots of 10 kg, 20 kg, 30 kg, 40 kg, 50 kg and 60 kg of cherries by square meter of drying area. The slowness of the drying led to the increasing of fungal development and ochratoxin A production in the cherries. Indeed, samples more contaminated were those from the lots of 50 kg and 60 kg of cherries by square meter of drying area with between 10% and 100% of infected beans and with levels of ochratoxin A ranging from 0.92 to 118.47 and 1.4 to 131.33 μg kg(-1) respectively. The slowness of the drying led also to the acidification of the cherries (pH=5.55-4.54) and the degradation of their chlorogenic acids content (13.03-11.69) while for their caffeine content (2.52-2.54), any significant difference was observed whatever the drying duration. Copyright © 2012 Elsevier Ltd. All rights reserved.

Effect of environmental factors on Fusarium population and associated trichothecenes in wheat grain grown in Jiangsu province, China.

PubMed

Dong, Fei; Qiu, Jianbo; Xu, Jianhong; Yu, Mingzheng; Wang, Shufang; Sun, Yue; Zhang, Gufeng; Shi, Jianrong

2016-08-02

The present study was performed to identify prevailing Fusarium species and the environmental factors affecting their frequencies and the contamination of grain with major mycotoxins in Jiangsu province. The precipitation levels were 184.2mm, 156.4mm, and 245.8mm in the years 2013-2015, respectively, and the temperature fluctuated by an average of 10.6±7.2°C in 2013, 10.9±7.2°C in 2014, and 10.6±6.3°C in 2015. Co-occurrence of deoxynivalenol (DON), 3-acetyldeoxynivalenol (3ADON), and 15-acetyldeoxynivalenol (15ADON) were observed in wheat. The average concentrations of DON were 879.3±1127.8, 627.8±640.5, and 1628.6±2,168.0μg/kg in 2013-2015, respectively. The average concentrations of 3ADON were 43.5±59.0, 71.2±102.5, and 33.5±111.9μg/kg in 2013-2015, respectively. We found that the average concentration of DON in wheat was positively correlated with precipitation (r=0.998, p<0.01), and 3ADON was negatively correlated with precipitation (r=-0.887, p<0.05). However, there was no correlation between precipitation and 15ADON or nivalenol (NIV). The differences in temperature were not as significant as the differences in rainfall amount over a short time period. Therefore, there were no correlations between temperature and the concentrations of trichothecenes, excluding 3ADON (r=0.996, p<0.01). Our data indicated that Fusarium asiaticum is the primary pathogenic fungus prevalent in the Fusarium head blight disease nursery. The trichothecene chemotype composition differed between Fusarium graminearum sensu stricto (s. str.) and F. asiaticum isolates. The 3ADON chemotype was found only among strains of F. asiaticum. The NIV chemotype was not observed among strains of F. graminearum, while the 15ADON chemotype represented 100% of the F. graminearum strains collected. The results of this study indicated no correlations between environmental conditions and the species or genetic chemotype composition of pathogens in Jiangsu province in 2013-2015. Copyright �

Stability of ochratoxin A (OTA) during processing and decaffeination in commercial roasted coffee beans.

PubMed

Nehad, E A; Farag, M M; Kawther, M S; Abdel-Samed, A K M; Naguib, K

2005-08-01

The fate of ochratoxin A (OTA) during the processing of artificially contaminated green coffee beans, the effect of decaffeination on the production of OTA in green and roasted coffee beans, and the effect of caffeine on the growth and OTA production by Aspergillus ochraceus were studied. The data indicated that the roasting, milling and decoction (brewing and Turkish coffee making) processes caused different percentage reductions in OTA. Decaffeinated samples showed a significantly higher concentration of OTA production than the caffeinated ones. A significantly higher percentage of OTA was reduced when the decaffeination process was performed before roasting treatment. Caffeine at 1.0 and 2.0% concentrations completely prevented OTA production and completely inhibited A. ochraceus growth in YES medium after 3-21 days.

Natural occurrence of ochratoxin A in wolfberry fruit wine marketed in China.

PubMed

Kuang, Ying; Qiu, Feng; Kong, Weijun; Yang, Meihua

2012-01-01

Wolfberry fruit wine (WFW) is widely used as a global functional food to improve the immune system and prevent human disease. A total of 36 bottled WFWs were randomly collected in China between 2005 and 2010. Samples were analysed for the presence of ochratoxin A (OTA) using immunoaffinity column (IAC) clean-up and high-performance liquid chromatography with fluorescence detection (HPLC-FLD). Positive results were confirmed by liquid chromatography-electrospray ionisation-tandem mass spectrometry (LC-ESI-MS/MS). The limit of detection (LOD), based on a signal-to-noise ratio of 3, was 0.05 ng mL⁻¹. Recoveries ranged from 78.3% to 94.7% and relative standard deviations from 1.1% to 4.3% within the spiking range of 0.2-20 ng mL⁻¹. OTA was detected in one sample, below the maximum allowable limit as established by the European community.

Biodegradation of Ochratoxin A for Food and Feed Decontamination

PubMed Central

Abrunhosa, Luís; Paterson, Robert R. M.; Venâncio, Armando

2010-01-01

Ochratoxin A (OTA) is one of the most important mycotoxins that is found in food and feed products. It has proven toxic properties, being primarily known for its nephrotoxicity and carcinogenicity to certain animal species. OTA is produced by several species of Aspergillus and Penicillium that can be found in a wide variety of agricultural products, which makes the presence of OTA in these products common. Many countries have statutory limits for OTA, and concentrations need to be reduced to as low as technologically possible in food and feed. The most important measures to be taken to control OTA are preventive in order to avoid fungal growth and OTA production. However, these measures are difficult to implement in all cases with the consequence of OTA remaining in agricultural commodities. Remediation processes are often used to eliminate, reduce or avoid the toxic effects of OTA. Biological methods have been considered increasingly as an alternative to physical and chemical treatments. However, examples of practical applications are infrequent. This review will focus on the (i) known microorganisms and enzymes that are able to biodegrade OTA; (ii) mode of action of biodegradation and (iii) current applications. A critical discussion about the technical applicability of these strategies is presented. PMID:22069627

Transfer of Ochratoxin A into Tea and Coffee Beverages

PubMed Central

Malir, Frantisek; Ostry, Vladimir; Pfohl-Leszkowicz, Annie; Toman, Jakub; Bazin, Ingrid; Roubal, Tomas

2014-01-01

Ochratoxin A (OTA) is nephrotoxic, hepatotoxic, immunotoxic, neurotoxic, reprotoxic, teratogenic, and carcinogenic (group 2B), being characterized by species and sex differences in sensitivity. Despite the fact that OTA is in some aspects a controversial topic, OTA is the most powerful renal carcinogen. The aim of this study was to make a small survey concerning OTA content in black tea, fruit tea, and ground roasted coffee, and to assess OTA transfer into beverages. OTA content was measured using a validated and accredited HPLC-FLD method with a limit of quantification (LOQ) of 0.35 ng/g. The OTA amount ranged from LOQ up to 250 ng/g in black tea and up to 104 ng/g in fruit tea. Black tea and fruit tea, naturally contaminated, were used to prepare tea infusions. The transfer from black tea to the infusion was 34.8% ± 1.3% and from fruit tea 4.1% ± 0.2%. Ground roasted coffee naturally contaminated at 0.92 ng/g was used to prepare seven kinds of coffee beverages. Depending on the type of process used, OTA transfer into coffee ranged from 22.3% to 66.1%. OTA intakes from fruit and black tea or coffee represent a non-negligible human source. PMID:25525684

Variation in the Microbiome, Trichothecenes, and Aflatoxins in Stored Wheat Grains in Wuhan, China.

PubMed

Yuan, Qing-Song; Yang, Peng; Wu, Ai-Bo; Zuo, Dong-Yun; He, Wei-Jie; Guo, Mao-Wei; Huang, Tao; Li, He-Ping; Liao, Yu-Cai

2018-04-24

Contamination by fungal and bacterial species and their metabolites can affect grain quality and health of wheat consumers. In this study, sequence analyses of conserved DNA regions of fungi and bacteria combined with determination of trichothecenes and aflatoxins revealed the microbiome and mycotoxins of wheat from different silo positions (top, middle, and bottom) and storage times (3, 6, 9, and 12 months). The fungal community in wheat on the first day of storage (T₀) included 105 classified species (81 genera) and 41 unclassified species. Four species had over 10% of the relative abundance: Alternaria alternata (12%), Filobasidium floriforme (27%), Fusarium graminearum (12%), and Wallemia sebi (12%). Fungal diversity and relative abundance of Fusarium in wheat from top silo positions were significantly lower than at other silo positions during storage. Nivalenol and deoxynivalenol in wheat were 13⁻34% higher in all positions at 3 months compared to T₀, and mycotoxins in wheat from middle and bottom positions at 6 to 12 months were 24⁻57% higher than at T₀. The relative abundance of toxigenic Aspergillus and aflatoxins were low at T₀ and during storage. This study provides information on implementation and design of fungus and mycotoxin management strategies as well as prediction models.

Variation in the Microbiome, Trichothecenes, and Aflatoxins in Stored Wheat Grains in Wuhan, China

PubMed Central

Yuan, Qing-Song; Yang, Peng; Zuo, Dong-Yun; He, Wei-Jie; Guo, Mao-Wei; Huang, Tao; Li, He-Ping; Liao, Yu-Cai

2018-01-01

Contamination by fungal and bacterial species and their metabolites can affect grain quality and health of wheat consumers. In this study, sequence analyses of conserved DNA regions of fungi and bacteria combined with determination of trichothecenes and aflatoxins revealed the microbiome and mycotoxins of wheat from different silo positions (top, middle, and bottom) and storage times (3, 6, 9, and 12 months). The fungal community in wheat on the first day of storage (T0) included 105 classified species (81 genera) and 41 unclassified species. Four species had over 10% of the relative abundance: Alternaria alternata (12%), Filobasidium floriforme (27%), Fusarium graminearum (12%), and Wallemia sebi (12%). Fungal diversity and relative abundance of Fusarium in wheat from top silo positions were significantly lower than at other silo positions during storage. Nivalenol and deoxynivalenol in wheat were 13–34% higher in all positions at 3 months compared to T0, and mycotoxins in wheat from middle and bottom positions at 6 to 12 months were 24–57% higher than at T0. The relative abundance of toxigenic Aspergillus and aflatoxins were low at T0 and during storage. This study provides information on implementation and design of fungus and mycotoxin management strategies as well as prediction models. PMID:29695035

Ochratoxin A production in relation to ecophysiological factors by Aspergillus section Nigri strains isolated from different substrates in Argentina.

PubMed

Astoreca, A; Magnoli, C; Barberis, C; Chiacchiera, S M; Combina, M; Dalcero, A

2007-12-15

Contamination of foodstuff with mycotoxins such as ochratoxins is a major matter of concern for human and animal health. In Aspergillus species, ochratoxin synthesis depends on several environmental factors. The aims of this work were to evaluate the effect of water activity (0.995-0.85), temperature (15, 25 and 30 degrees C), incubation time (7, 14 and 21 days) and their interactions on OTA production on peanut, maize kernels, dried grapes and coffee beans meal extract agar medium by eight strains of Aspergillus section Nigri isolated from human food in Argentina. The optimum temperature for OTA production was 25 or 30 degrees C depending on the strains assayed, in most cases the highest OTA levels were achieved after 7 days of incubation, whereas this situation occurred at 15 degrees C after 14 days of incubation for only one strain. The maximum OTA level was obtained at earlier growth states when incubation temperature increased. In general, OTA concentration increased as water activity (a(W)) increased with no significant production at 0.85-0.91 a(W) under all temperature levels tested. Production occurred over a range of temperatures (15-30 degrees C) with optimum production at 30 degrees C depending on a(W) assayed. The knowledge of Aspergillus section Nigri ecophysiology is critical in the development and prediction of the risk models of raw material and final product contamination by these species under fluctuating and interacting environmental parameters.

Early activation of wheat polyamine biosynthesis during Fusarium head blight implicates putrescine as an inducer of trichothecene mycotoxin production

PubMed Central

2010-01-01

Background The fungal pathogen Fusarium graminearum causes Fusarium Head Blight (FHB) disease on wheat which can lead to trichothecene mycotoxin (e.g. deoxynivalenol, DON) contamination of grain, harmful to mammalian health. DON is produced at low levels under standard culture conditions when compared to plant infection but specific polyamines (e.g. putrescine and agmatine) and amino acids (e.g. arginine and ornithine) are potent inducers of DON by F. graminearum in axenic culture. Currently, host factors that promote mycotoxin synthesis during FHB are unknown, but plant derived polyamines could contribute to DON induction in infected heads. However, the temporal and spatial accumulation of polyamines and amino acids in relation to that of DON has not been studied. Results Following inoculation of susceptible wheat heads by F. graminearum, DON accumulation was detected at two days after inoculation. The accumulation of putrescine was detected as early as one day following inoculation while arginine and cadaverine were also produced at three and four days post-inoculation. Transcripts of ornithine decarboxylase (ODC) and arginine decarboxylase (ADC), two key biosynthetic enzymes for putrescine biosynthesis, were also strongly induced in heads at two days after inoculation. These results indicated that elicitation of the polyamine biosynthetic pathway is an early response to FHB. Transcripts for genes encoding enzymes acting upstream in the polyamine biosynthetic pathway as well as those of ODC and ADC, and putrescine levels were also induced in the rachis, a flower organ supporting DON production and an important route for pathogen colonisation during FHB. A survey of 24 wheat genotypes with varying responses to FHB showed putrescine induction is a general response to inoculation and no correlation was observed between the accumulation of putrescine and infection or DON accumulation. Conclusions The activation of the polyamine biosynthetic pathway and putrescine in

Rapid analysis of aflatoxins B1, B2, and ochratoxin A in rice samples using dispersive liquid-liquid microextraction combined with HPLC.

PubMed

Lai, Xian-Wen; Sun, Dai-Li; Ruan, Chun-Qiang; Zhang, He; Liu, Cheng-Lan

2014-01-01

A novel, simple, and rapid method is presented for the analysis of aflatoxin B1, aflatoxin B2, and ochratoxin A in rice samples by dispersive liquid-liquid microextraction combined with LC and fluorescence detection. After extraction of the rice samples with a mixture of acetonitrile/water/acetic acid, mycotoxins were rapidly partitioned into a small volume of organic solvent (chloroform) by dispersive liquid-liquid microextraction. The three mycotoxins were simultaneously determined by LC with fluorescence detection after precolumn derivatization for aflatoxin B1 and B2. Parameters affecting both extraction and dispersive liquid-liquid microextraction procedures, including the extraction solvent, the type and volume of extractant, the volume of dispersive solvent, the addition of salt, the pH and the extraction time, were optimized. The optimized protocol provided an enrichment factor of approximately 1.25 and with detection of limits (0.06-0.5 μg/kg) below the maximum levels imposed by current regulations for aflatoxins and ochratoxin A. The mean recovery of three mycotoxins ranged from 82.9-112%, with a RSD less than 7.9% in all cases. The method was successfully applied to measure mycotoxins in commercial rice samples collected from local supermarkets in China. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Determination of type A trichothecenes in coix seed by magnetic solid-phase extraction based on magnetic multi-walled carbon nanotubes coupled with ultra-high performance liquid chromatography-tandem mass spectrometry.

PubMed

Dong, Maofeng; Si, Wenshuai; Wang, Weimin; Bai, Bing; Nie, Dongxia; Song, Weiguo; Zhao, Zhihui; Guo, Yirong; Han, Zheng

2016-09-01

Magnetic solid-phase extraction (m-SPE) is a promising sample preparation approach due to its convenience, speed, and simplicity. For the first time, a rapid and reliable m-SPE approach using magnetic multi-walled carbon nanotubes (m-MWCNTs) as the adsorbent was proposed for purification of type A trichothecenes including T-2 toxins (T2), HT-2 toxins (HT-2), diacetoxyscirpenol (DAS), and neosolaniol (NEO) in coix seed. The m-MWCNTs were synthesized by assembling the magnetic nanoparticles (Fe3O4) with MWCNTs by sonication through an aggregation wrap mechanism, and characterized by transmission electron microscope. Several key parameters affecting the performance of the procedure were extensively investigated including extraction solutions, desorption solvents, and m-MWCNT amounts. Under the optimal sample preparation conditions followed by analysis with ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS), high sensitivity (limit of quantification in the range of 0.3-1.5 μg kg(-1)), good linearity (R (2) > 0.99), satisfactory recovery (73.6-90.6 %), and acceptable precision (≤2.5 %) were obtained. The analytical performance of the developed method has also been successfully evaluated in real coix seed samples. Graphical Abstract Flow chart of determination of type A trichothecenes in coix seed by magnetic solid-phase extraction coupled with ultra-high performance liquid chromatography-tandem mass spectrometry.

Phylogenetic evidence for an animal pathogen origin of ergot and the grass endophytes.

PubMed

Spatafora, J W; Sung, G-H; Sung, J-M; Hywel-Jones, N L; White, J F

2007-04-01

Grass-associated fungi (grass symbionts) in the family Clavicipitaceae (Ascomycota, Hypocreales) are species whose host range is restricted to the plant family Poaceae and rarely Cyperaceae. The best-characterized species include Claviceps purpurea (ergot of rye) and Neotyphodium coenophialum (endophyte of tall fescue). They have been the focus of considerable research due to their importance in agricultural and grassland ecosystems and the diversity of their bioactive secondary metabolites. Here we show through multigene phylogenetic analyses and ancestral character state reconstruction that the grass symbionts in Clavicipitaceae are a derived group that originated from an animal pathogen through a dynamic process of interkingdom host jumping. The closest relatives of the grass symbionts include the genera Hypocrella, a pathogen of scale insects and white flies, and Metarhizium, a generalist arthropod pathogen. These data do not support the monophyly of Clavicipitaceae, but place it as part of a larger clade that includes Hypocreaceae, a family that contains mainly parasites of other fungi. A minimum of 5-8 independent and unidirectional interkingdom host jumps has occurred among clavicipitaceous fungi, including 3-5 to fungi, 1-2 to animals, and 1 to plants. These findings provide a new evolutionary context for studying the biology of the grass symbionts, their role in plant ecology, and the evolution of host affiliation in fungal symbioses.

Transdermal Patch of Rotigotine Attenuates Freezing of Gait in Patients with Parkinson's Disease: An Open-Label Comparative Study of Three Non-Ergot Dopamine Receptor Agonists.

PubMed

Ikeda, Ken; Hirayama, Takehisa; Takazawa, Takanori; Kawabe, Kiyokazu; Iwasaki, Yasuo

Objective Parkinson's disease (PD) is characterized by the progressive degeneration of the nigrostriatal dopaminergic neurons. Rotigotine is a non-ergot dopamine receptor agonist (DA). Its transdermal patch maintains the effective concentrations for 24 hours. Freezing of gait (FOG) is a common and devastating symptom in PD patients. Little is known about therapeutic effects of rotigotine on FOG in PD patients. Herein we compared how three non-ergot DAs of rotigotine, pramipexole LA and ropinirole CR influence FOG, besides classical motor deficits in PD patients. Methods Rotigotine (maintenance doses of 9-27 mg/day) was administered in 51 patients, 36 patients received pramipexole LA (1.5-4.5 mg/day) and 35 patients received ropinirole CR (8-16 mg/day). The Unified PD Rating Scale (UPDRS) parts I-IV, FOG questionnaire (16 items) and wearing off time were examined from baseline to 7 months after DA administration. UPDRS parts I-IV were evaluated during on time and FOG was recorded during off time if patients experienced wearing off. Results A total of 111 patients completed the study. UPDRS parts II-III scores and wearing off time were significantly reduced after each DA treatment compared to baseline. FOG was found in 54 patients (49%). Most patients developed FOG during off time only. FOG scores were significantly decreased at 2 months after rotigotine treatment whereas pramipexole LA and ropinirole treatment did not alter FOG scores. Conclusion The present study indicates that transdermal patch of rotigotine attenuated the FOG off time. The similar binding affinities to dopamine receptors between rotigotine and dopamine, and 24 hours steady hemodynamics could contribute to the therapeutic mechanism of rotigotine on FOG in PD patients with wearing off.

Occurrence of Ochratoxin A in Grape Juice of Iran

PubMed Central

Yusefi, Jamal; Valaee, Mehran; Nazari, Firouzeh; Maleki, Javad; Mottaghianpour, Ehsan; Khosrokhavar, Roya; Hosseini, Mir-Jamal

2018-01-01

Ochratoxin A (OTA) is one of the most important mycotoxins that contaminate a broad range of agricultural and food products. In this study, the occurrence of OTA in available brands of grape juice in Iran purchased from retail outlets or producer were determined for the first time using high-performance liquid chromatography (HPLC) with immunoaffinity columns(IAC) as the clean-up step. The average recoveries for OTA in grape juice ranged from 54.2 to 86.6% with the coefficient of variation lower than 17.3% in lowest spiked level (0.5 µg/L). The estimated LOD and LOQ of OTA were 0.04 µg/L and 0.125 µg/L, respectively. In our study, 70 samples of grape juice evaluated for OTA content. The results showed that in 39 out of 70 samples (55.7%) OTA levels were above the LOQ with the maximum level of 2.6 µg/L and the mean contamination was 0.5 µg/L. Although the mean contamination of OTA in analyzing samples was lower than the MRL set by EU, the high incidence of contamination in these products is worried. Considering the importance of OTA in public health, control of pre- and post-harvest, storage and grape juice manufacturing process, such as HACCP, GAP, and GMP recommended preventive measures are required. PMID:29755546

Fungi producing significant mycotoxins.

PubMed

2012-01-01

Mycotoxins are secondary metabolites of microfungi that are known to cause sickness or death in humans or animals. Although many such toxic metabolites are known, it is generally agreed that only a few are significant in causing disease: aflatoxins, fumonisins, ochratoxin A, deoxynivalenol, zearalenone, and ergot alkaloids. These toxins are produced by just a few species from the common genera Aspergillus, Penicillium, Fusarium, and Claviceps. All Aspergillus and Penicillium species either are commensals, growing in crops without obvious signs of pathogenicity, or invade crops after harvest and produce toxins during drying and storage. In contrast, the important Fusarium and Claviceps species infect crops before harvest. The most important Aspergillus species, occurring in warmer climates, are A. flavus and A. parasiticus, which produce aflatoxins in maize, groundnuts, tree nuts, and, less frequently, other commodities. The main ochratoxin A producers, A. ochraceus and A. carbonarius, commonly occur in grapes, dried vine fruits, wine, and coffee. Penicillium verrucosum also produces ochratoxin A but occurs only in cool temperate climates, where it infects small grains. F. verticillioides is ubiquitous in maize, with an endophytic nature, and produces fumonisins, which are generally more prevalent when crops are under drought stress or suffer excessive insect damage. It has recently been shown that Aspergillus niger also produces fumonisins, and several commodities may be affected. F. graminearum, which is the major producer of deoxynivalenol and zearalenone, is pathogenic on maize, wheat, and barley and produces these toxins whenever it infects these grains before harvest. Also included is a short section on Claviceps purpurea, which produces sclerotia among the seeds in grasses, including wheat, barley, and triticale. The main thrust of the chapter contains information on the identification of these fungi and their morphological characteristics, as well as factors

Post-harvest practices linked with ochratoxin A contamination of coffee in three provinces of Cordillera Administrative Region, Philippines.

PubMed

Barcelo, Jonathan M; Barcelo, Racquel C

2018-02-01

One of the emerging concerns in the Cordillera Administrative Region, Philippines is ochratoxin A (OTA) contamination in coffee. During 2015 to 2016, a total of 51 Arabica (Coffea arabica) coffee samples from Benguet province and 71 Robusta (Coffea canephora var. Robusta) coffee samples from the provinces of Ifugao and Kalinga were analysed for OTA contamination. The OTA-producing fungal contaminants during drying and storage of Arabica and Robusta coffee were Aspergillus niger and Aspergillus ochraceus. Ochratoxin A was more commonly detected in Robusta coffee (36.6%) than in Arabica coffee (21.6%). Among the contaminated samples, Robusta coffee cherries in the drying yard had the highest mean OTA level (120.2 μg kg -1 , n = 10) while roasted Robusta coffee beans had the lowest mean level (4.8 μg kg -1 , n = 9). The onset of contamination of Arabica coffee occurred during storage, with a mean OTA level of 46.7 μg kg -1 (n = 9). Roasted coffee had lower OTA content although five samples had levels >5.0 μg kg -1 . Pearson Chi-square analysis (χ 2 ) and Fisher's exact test revealed that several post-harvest practices involving non-removal of the husk or hull and mixing of defective coffee were significantly associated with the occurrence of OTA during drying and storage (p < 0.05). No significant associations, however, were identified during roasting. This study suggests that the post-harvest practices in Cordillera Administrative Region should focus on the removal of defective coffee in all stages of post-harvest and rapid reduction of moisture content particularly during drying.

Comparative Toxicokinetics and Plasma Protein Binding of Ochratoxin A in Four Avian Species.

PubMed

Devreese, Mathias; Croubels, Siska; De Baere, Siegrid; Gehring, Ronette; Antonissen, Gunther

2018-03-07

Ochratoxin A (OTA, 0.25 mg/kg body weight) was absorbed rapidly ( T max = 0.31-1.88 h) in all avian species (broiler chickens, laying hens, turkeys, and Muscovy ducks) but more slowly in broiler chickens ( T max = 1.43-4.63 h). The absolute oral bioavailability was complete in these bird species (88.0-109.6%). Ducks have a significantly higher volume of distribution ( V d ) and turkeys a lower V d compared to chickens and layers (broiler chickens, 0.27 ± 0.12 L/kg; layers, 0.23 ± 0.08 L/kg; turkeys, 0.18 ± 0.04 L/kg; ducks, 0.76 ± 0.44 L/kg). This difference in V d can be attributed to the species-dependent differences in plasma protein binding of OTA, namely ranging between 82.2 and 88.9% in ducks and between 96.5 and 98.8% in turkeys. No significant gender differences were found in toxicokinetics or plasma protein binding.

Aflatoxins and ochratoxin a reduction in black and white pepper by gamma radiation

NASA Astrophysics Data System (ADS)

Jalili, M.; Jinap, S.; Noranizan, M. A.

2012-11-01

Irradiation is an important means of decontamination of food commodities, especially spices. The aim of the current study was to investigate the efficacy of gamma radiation (60Co) for decontaminating ochratoxin A (OTA) and aflatoxins B1 (AFB1), B2 (AFB2), G1 (AFG1) and G2 (AFG2) residues in artificially contaminated black and white pepper samples. The moisture content of the pepper samples was set at 12% or 18%, and the applied gamma dose ranged from 5 to 30 kGy. Mycotoxin levels were determined by high-performance liquid chromatography (HPLC) after immunoaffinity column (IAC) chromatography. Both the gamma irradiation dose and moisture content showed significant effects (P<0.05) on mycotoxin reduction. The maximum toxin reductions, found at 18% moisture content and 30 kGy, were 55.2%, 50.6%, 39.2%, 47.7% and 42.9% for OTA, AFB1, AFB2, AFG1 and AFG2, respectively.

Producers and Important Dietary Sources of Ochratoxin A and Citrinin

PubMed Central

Ostry, Vladimir; Malir, Frantisek; Ruprich, Jiri

2013-01-01

Ochratoxin A (OTA) is a very important mycotoxin, and its research is focused right now on the new findings of OTA, like being a complete carcinogen, information about OTA producers and new exposure sources of OTA. Citrinin (CIT) is another important mycotoxin, too, and its research turns towards nephrotoxicity. Both additive and synergistic effects have been described in combination with OTA. OTA is produced in foodstuffs by Aspergillus Section Circumdati (Aspergillus ochraceus, A. westerdijkiae, A. steynii) and Aspergillus Section Nigri (Aspergillus carbonarius, A. foetidus, A. lacticoffeatus, A. niger, A. sclerotioniger, A. tubingensis), mostly in subtropical and tropical areas. OTA is produced in foodstuffs by Penicillium verrucosum and P. nordicum, notably in temperate and colder zones. CIT is produced in foodstuffs by Monascus species (Monascus purpureus, M. ruber) and Penicillium species (Penicillium citrinum, P. expansum, P. radicicola, P. verrucosum). OTA was frequently found in foodstuffs of both plant origin (e.g., cereal products, coffee, vegetable, liquorice, raisins, wine) and animal origin (e.g., pork/poultry). CIT was also found in foodstuffs of vegetable origin (e.g., cereals, pomaceous fruits, black olive, roasted nuts, spices), food supplements based on rice fermented with red microfungi Monascus purpureus and in foodstuffs of animal origin (e.g., cheese). PMID:24048364

Mycobiota and concentration of ochratoxin A in concentrated poultry feed from Venezuela.

PubMed

Figueroa, S; Centeno, S; Calvo, M A; Rengel, A; Adelantado, C

2009-04-01

The main objective of this study was to evaluate the frequency distribution of mycobiota and the concentration of Ochratoxin A (OTA) in 50 samples from one company of commercial brand poultry feed produced in Venezuela. The concentration of OTA in the samples analyzed was determined using the competitive ELISA method. The most frequently isolated genera of moulds were Aspergillus (36%) and Penicillium (20%). Of these genera, the most frequently isolated species were Aspergillus flavus, Aspergillus terreus and Penicillium citrinum. Ochratoxigenic species such as Eurotium herbariorum, Aspergillus niger, Aspergillus ochraceus and Aspergillus glaucus, were also found with lower frequency. Rhodotorula mucilaginosa was the only yeast isolated. 94% of the samples presented contamination by OTA in a range between 2.558 and 31.978 microg kg(-1) feed and 42% of them presented OTA levels from 10 up to 20 microg kg(-1). The findings of this investigation show that 84% of the samples of concentrated feed for meat poultry surpass the maximum permitted limit for OTA of 5 microg kg(-1), established in the majority of countries in which regulations are placed.

Screening and Identification of Novel Ochratoxin A-Producing Fungi from Grapes

PubMed Central

Zhang, Xiaoyun; Li, Yulin; Wang, Haiying; Gu, Xiangyu; Zheng, Xiangfeng; Wang, Yun; Diao, Junwei; Peng, Yaping; Zhang, Hongyin

2016-01-01

Ochratoxin A (OTA) contamination has been established as a world-wide problem. In this study, the strains with the ability of OTA production were screened by analyzing the green fluorescence of the isolates colonies from the grapes in Zhenjiang with 365 nm UV light and confirmed by HPLC with fluorescent detection (HPLC-FLD). The results showed that seven isolates acquired the characteristic of the fluorescence, of which only five showed the ability of OTA production as confirmed by HPLC-FLD analysis. The five OTA-producing strains were identified based on comparative sequence analysis of three conserved genes (ITS, BenA and RPB2) of the strains, and they are Talaromyces rugulosus (O1 and Q3), Penicillium commune (V5-1), Penicillium rubens (MQ-5) and Aspergillus aculeatus (MB1-1). There are two Penicillium species of the five OTA-producing strains and our study is the first to report that P. rubens, T. rugulosus and A. aculeatus can produce OTA. This work would contribute to comprehensively understanding the fungi with an OTA-producing ability in grapes before harvest and then take effective measures to prevent OTA production. PMID:27845758

Exposure assessment to ochratoxin A in Chinese wine.

PubMed

Zhong, Qi Ding; Li, Guo Hui; Wang, Dao Bing; Shao, Yi; Li, Jing Guang; Xiong, Zheng He; Wu, Yong Ning

2014-09-03

A rapid, sensitive, reproducible, and inexpensive method of high-performance liquid chromatography with fluorescence detection after an anion-exchange solid-phase extraction cleanup step for the analysis of ochratoxin A (OTA) in Chinese wine was developed. The average recovery rate and the average RSD of recovery were 97.47% and about 4%. The relative standard deviations of both the interday and intraday precision were 6.7 and 12.6%, respectively. The limits of detection and quantitation were determined to be 0.01 and 0.03 μg/L, respectively. A total of 223 samples from the major wine-producing areas of China were analyzed for OTA. OTA was detected at levels of 0.01-0.98 μg/L. The mean was 0.15 μg/L. Then, participants as representative inhabitants were invited to answer the designed questionnaire about the quantity and frequency of wine consumption. All data were simulated by the point evaluation for the risk assessment of OTA contamination from wine. Those results indicated that daily intake (DI) of OTA for the average adult consumer varies between 0.86 and 1.08 ng/kg bw per week, which was lower than all the reference standards. However, the DI value (4.38-5.54 ng/kg bw per week) in the high percentile (97.5) was slightly above 5% PTWI (100 ng kg(-1) week(-1)) of the JECFA. In conclusion, OTA exposure from Chinese wine has no risk of harm. This research will provide the scientific basis for determining the maximum limit of OTA content in Chinese wine.

LC-MS/MS multi-method for mycotoxins after single extraction, with validation data for peanut, pistachio, wheat, maize, cornflakes, raisins and figs.

PubMed

Spanjer, Martien C; Rensen, Peter M; Scholten, Jos M

2008-04-01

, raisins and figs. The method has been compared with the official EU method for the determination of aflatoxins in food and relevant FAPAS rounds. The multi-mycotoxin method has been proven by the detection of more than one mycotoxin in maize, buckwheat, figs and nuts. The LC-MS/MS technique has also been applied to baby food, which is subject to lower limits for aflatoxin B1 and ochratoxin A, ergot alkaloids in naturally contaminated rye and freeze-dried silage samples.

Further aspects of ochratoxin A-cation interactions: complex formation with zinc ions and a novel analytical application of ochratoxin A-magnesium interaction in the HPLC-FLD system.

PubMed

Poór, Miklós; Kuzma, Mónika; Matisz, Gergely; Li, Yin; Perjési, Pál; Kunsági-Máté, Sándor; Kőszegi, Tamás

2014-04-10

Ochratoxin A (OTA) is a mycotoxin produced by different Aspergillus and Penicillium species. Since its mechanism of action is not fully understood yet, it is important to gain further insight into different interactions of OTA at the molecular level. OTA is found worldwide in many foods and drinks. Moreover, it can also be detected in human and animal tissues and body fluids, as well. Therefore, the development of highly sensitive quantitative methods for the determination of OTA is of utmost importance. OTA most likely forms complexes with divalent cations, both in cells and body fluids. In the present study, the OTA-zinc interaction was investigated and compared to OTA-magnesium complex formation using fluorescence spectroscopy and molecular modeling. Our results show that zinc(II) ion forms a two-fold higher stable complex with OTA than magnesium(II) ion. In addition, based on the enhanced fluorescence emission of OTA in its magnesium-bound form, a novel RP-HPLC-fluorescence detector (FLD) method was also established. Our results highlight that the application of magnesium chloride in alkaline eluents results in an approximately two-fold increase in sensitivity using the HPLC-FLD technique.

Transdermal Patch of Rotigotine Attenuates Freezing of Gait in Patients with Parkinson's Disease: An Open-Label Comparative Study of Three Non-Ergot Dopamine Receptor Agonists

PubMed Central

Ikeda, Ken; Hirayama, Takehisa; Takazawa, Takanori; Kawabe, Kiyokazu; Iwasaki, Yasuo

2016-01-01

Objective Parkinson's disease (PD) is characterized by the progressive degeneration of the nigrostriatal dopaminergic neurons. Rotigotine is a non-ergot dopamine receptor agonist (DA). Its transdermal patch maintains the effective concentrations for 24 hours. Freezing of gait (FOG) is a common and devastating symptom in PD patients. Little is known about therapeutic effects of rotigotine on FOG in PD patients. Herein we compared how three non-ergot DAs of rotigotine, pramipexole LA and ropinirole CR influence FOG, besides classical motor deficits in PD patients. Methods Rotigotine (maintenance doses of 9-27 mg/day) was administered in 51 patients, 36 patients received pramipexole LA (1.5-4.5 mg/day) and 35 patients received ropinirole CR (8-16 mg/day). The Unified PD Rating Scale (UPDRS) parts I-IV, FOG questionnaire (16 items) and wearing off time were examined from baseline to 7 months after DA administration. UPDRS parts I-IV were evaluated during on time and FOG was recorded during off time if patients experienced wearing off. Results A total of 111 patients completed the study. UPDRS parts II-III scores and wearing off time were significantly reduced after each DA treatment compared to baseline. FOG was found in 54 patients (49%). Most patients developed FOG during off time only. FOG scores were significantly decreased at 2 months after rotigotine treatment whereas pramipexole LA and ropinirole treatment did not alter FOG scores. Conclusion The present study indicates that transdermal patch of rotigotine attenuated the FOG off time. The similar binding affinities to dopamine receptors between rotigotine and dopamine, and 24 hours steady hemodynamics could contribute to the therapeutic mechanism of rotigotine on FOG in PD patients with wearing off. PMID:27725534

Two-dimensional MoS2 as a nano-binder for ssDNA: Ultrasensitive aptamer based amperometric detection of Ochratoxin A.

PubMed

Tang, Juan; Huang, Yapei; Cheng, Yu; Huang, Lulu; Zhuang, Junyang; Tang, Dianping

2018-02-07

Two-dimensional (2D) MoS 2 is found to possess different affinities for ssDNA and dsDNA. This finding is exploited in an amperometric aptamer-based method for the determination of the mycotoxin ochratoxin A (OTA). Initially, a dsDNA probe (formatted through the hybridization of OTA-aptamer with an auxiliary DNA) is self-assembled on a gold electrode. Upon introduction of OTA, it will bind to the aptamer and cause the unwinding of dsDNA, while the auxiliary DNA (with single-stranded structure) remains on the electrode. Since the affinity of 2D MoS 2 for ssDNA is considerably larger than that for dsDNA, it will be adsorbed on the electrode by binding to the auxiliary DNA. Notably, 2D MoS 2 possesses peroxidase-like activity. Hence, it can catalyze the amplification of electrochemical signal of the hydroquinone/benzoquinone redox system. Under optimal conditions, the amperometric signal (best measured at -0.2 V vs. SCE) increases with increasing OTA concentration in the range from 0.5 pg·mL -1 to 1.0 ng·mL -1 , with a lower detection limit of 0.23 pg·mL -1 . The method was applied to the determination of OTA in spiked red wine. Graphical abstract Herein we construct a convenient electrochemical aptasensor for sensitive monitor of ochratoxin A by using 2D MoS 2 as a nano-binder to catalyze the amplification of electrochemical signal from hydroquinone/benzoquinone system.

Facile synthesis of Fe3O4/g-C3N4/HKUST-1 composites as a novel biosensor platform for ochratoxin A.

PubMed

Hu, Shuisheng; Ouyang, Wenjun; Guo, Longhua; Lin, Zhenyu; Jiang, Xiaohua; Qiu, Bin; Chen, Guonan

2017-06-15

A fluorescent biosensor for ochratoxin A was fabricated on the basis of a new nanocomposite (Fe 3 O 4 /g-C 3 N 4 /HKUST-1 composites). Fe 3 O 4 /g-C 3 N 4 /HKUST-1 was synthesized in this work for the first time, which combined HKUST-1 with g-C 3 N 4 to improve its chemical stability. Fe 3 O 4 /g-C 3 N 4 /HKUST-1 composites have strong adsorption capacity for dye-labeled aptamer and are able to completely quench the fluorescence of the dye through the photoinduced electron transfer (PET) mechanism. In the presence of ochratoxin A (OTA), it can bind with the aptamer with high affinity, causing the releasing of the dye-labeled aptamer from the Fe 3 O 4 /g-C 3 N 4 /HKUST-1 and therefore results in the recovery of fluorescence. The fluorescence intensity of the biosensor has a linear relationship with the OTA concentration in the range of 5.0-160.0ng/mL. The LOD of sensor is 2.57ng/mL (S/N=3). This fluorescence sensor based on the Fe 3 O 4 /g-C 3 N 4 /HKUST-1 composites has been applied to detect OTA in corn with satisfying results. Copyright © 2016. Published by Elsevier B.V.

1H NMR Spectroscopy-Based Metabolomic Assessment of Uremic Toxicity, with Toxicological Outcomes, in Male Rats Following an Acute, Mid-Life Insult from Ochratoxin A

PubMed Central

Mantle, Peter G.; Nicholls, Andrew W.; Shockcor, John P.

2011-01-01

Overt response to a single 6.25 mg dose of ochratoxin A (OTA) by oral gavage to 15 months male rats was progressive loss of weight during the following four days. Lost weight was restored within one month and animals had a normal life-span without OTA-related terminal disease. Decline in plasma OTA concentration only commenced four days after dosing, while urinary excretion of OTA and ochratoxin alpha was ongoing. During a temporary period of acute polyuria, a linear relationship between urine output and creatinine concentration persisted. Elimination of other common urinary solutes relative to creatinine was generally maintained during the polyuria phase, except that phosphate excretion increased temporarily. 1H NMR metabolomic analysis of urine revealed a progressive cyclic shift in the group principal components data cluster from before dosing, throughout the acute insult phase, and returning almost completely to normality when tested six months later. Renal insult by OTA was detected by 1H NMR within a day of dosing, as the most sensitive early indicator. Notable biomarkers were trimethylamine N-oxide and an aromatic urinary profile dominated by phenylacetylglycine. Tolerance of such a large acute insult by OTA, assessed by rat natural lifetime outcomes, adds a new dimension to toxicology of this xenobiotic. PMID:22069722

Postharvest production of ochratoxin A by Aspergillus ochraceus and Penicillium viridicatum in barley with different protein levels.

PubMed Central

Häggblom, P E; Ghosh, J

1985-01-01

The production of ochratoxin A (OA) in barley by Aspergillus ochraceus and Penicillium viridicatum was measured at 12 and 25 degrees C. The grain had been fertilized with various amounts of nitrogen fertilizer (0, 90, or 240 kg/ha) and contained (at crop maturity) 9.1, 10.4, or 12.0% protein, respectively. The production of OA by both fungi increased as the protein concentration increased. Glutamic acid and proline were enriched relative to other amino acids as the protein concentration increased. The differences in OA production could not be explained by a differential effect of protein or amino acids on fungal growth in barley. However, glutamic acid and proline enhanced OA production in liquid cultures of both A. ochraceus and P. viridicatum. PMID:4004212

FTIR-ATR infrared spectroscopy for the detection of ochratoxin A in dried vine fruit.

PubMed

Galvis-Sánchez, Andrea C; Barros, Antonio; Delgadillo, Ivonne

2007-11-01

A method of screening sultanas for ochratoxin A (OTA) contamination, using mid-infrared spectroscopy/Golden Gate single-reflection ATR (attenuated total reflection), is described. The main spectral characteristics of sultanas from different sources were identified in a preliminary acquisition and spectral analysis study. Principal component analysis (PCA) showed that samples of various origins had different spectral characteristics, especially in water content and the fingerprint region. A lack of reproducibility was observed in the spectra acquired on different days. However, spectral repeatability was greatly improved when water activity of the sample was set at 0.62. A calibration curve of OTA was constructed in the range 10-40 microg OTA kg(-1). Samples with OTA levels higher than 20 microg kg(-1) were separated from samples contaminated with a lower concentration (10 microg OTA kg(-1)) and from uncontaminated samples. The reported methodology is a reliable and simple technique for screening dried vine fruit for OTA.

Solvent comparison in the isolation, solubilization, and toxicity of Stachybotrys chartarum spore trichothecene mycotoxins in an established in vitro luminescence protein translation inhibition assay.

PubMed

Black, J A; Foarde, K K; Menetrez, M Y

2006-08-01

It is well known that non-viable mold contaminants such as macrocyclic trichothecene mycotoxins of Stachybotrys chartarum are highly toxinigenic to humans. However, the method of recovering native mycotoxin has been without consensus. Inconsistencies occur in the methods of isolation, suspension, preparation, and quantitation of the mycotoxin from the spores. The purpose of this study was to provide quantitatively comparative data on three concurrent preparations of 10(6)S. chartarum spores. The experiments were designed to specifically evaluate a novel method of mycotoxin extraction, solubilization, and the subsequent inhibitory effect in an established in vitro luminescence protein translation assay from 30 day-old spores. The mycotoxin-containing spores swabbed from wallboard cultures were milled with and without glass beads in 100% methanol, 95% ethanol, or water. Milled spore lysates were cleared of cell debris by filter centrifugation followed by a second centrifugation through a 5000 MWCO filter to remove interfering proteins and RNases. Cleared lysate was concentrated by centrivap and suspended in either alcohol or water as described. The suspensions were used immediately in the in vitro luminescence protein translation assay with the trichothecene, T-2 toxin, as a control. Although, mycotoxin is reported to be alcohol soluble, the level of translation inhibition was not reliably satisfactory for either the methanol or ethanol preparations. In fact, the methanol and ethanol control reactions were not significantly different than the alcohol prepared spore samples. In addition, we observed that increasing amounts of either alcohol inhibited the reaction in a dose dependent manner. This suggests that although alcohol isolation of mycotoxin is desirable in terms of time and labor, the presence of alcohol in the luminescence protein translation reaction was not acceptable. Conversely, water extraction of mycotoxin demonstrated a dose dependent response, and

Mycoflora and mycotoxin contamination of Roundup Ready soybean harvested in the Pampean Region, Argentina.

PubMed

Garrido, Carolina E; González, Héctor H L; Salas, María Paula; Resnik, Silvia L; Pacin, Ana M

2013-08-01

A total of 89 freshly harvested soybean seed samples (Roundup Ready [transgenic] soybean cultivars) from the 2010/2011 crop season were collected from five locations in the Northern Pampean Region II, Argentina. These samples were analyzed for internal mycoflora, toxin production of isolated fungi, and for a range of mycotoxins. Mycotoxin analysis of aflatoxins (AFs), zearalenone (ZEA), fumonisins (FBs) and ochratoxin A (OTA) was done by HPLC-FLD (high performance liquid chromatography with postcolumn fluorescence derivatization), alternariol and alternariol monomethyl ether with HPLC-UV (HPLC with UV detection), trichothecenes (deoxynivalenol, nivalenol, T-2 toxin, HT-2 toxin, fusarenon X, 3-acetyldeoxynivalenol and 15-acetyldeoxynivalenol were analyzed by GC-ECD (gas chromatography with electron capture detector). Fungal colonization was more frequently found for samples from América, Saladillo and Trenque Lauquen than for samples from General Villegas and Trenel; a total of 1,401 fungal isolates were obtained from the soybean seeds. The most commonly identified fungal genera were Alternaria, Sclerotinia, Chaetomium, Cladosporium, Aspergillus, Penicillium, Phomopsis and Fusarium. Alternaria alternata, A.tenuissima, Aspergillus flavus, Penicillium citrinum, Fusarium verticillioides and F.semitectum were the predominant toxigenic fungal species. Mycotoxin production was confirmed for several isolates of toxigenic species, including Aspergillus flavus, A. parasiticus, Alternaria alternata, A.tenuissima, Fusarium graminearum, F semitectum and F. verticillioides. In particular, the percentage of mycotoxigenic Alternaria alternata (100%), A.tenuissima (95%) and aflatoxigenic strains of A. flavus (57%) were remarkably high. Although none of the mycotoxins, AFs, ZEA, FBs, trichothecenes and OTA, were directly detected in samples of soybean seeds, the frequent presence of toxigenic fungal species indicates the risk of multiple mycotoxin contamination.

Ochratoxin A removal by Saccharomyces cerevisiae strains: effect of wine-related physicochemical factors.

PubMed

Petruzzi, Leonardo; Sinigaglia, Milena; Corbo, Maria Rosaria; Beneduce, Luciano; Bevilacqua, Antonio

2013-07-01

This study investigated the effect of some physicochemical parameters on the removal of ochratoxin A (OTA) by yeasts. Two wild strains of Saccharomyces cerevisiae (W47 and Y28) were used to assess OTA removal under various conditions of temperature, pH, ethanol content and incubation time. All samples were analysed for OTA concentration by enzyme-linked immunosorbent assay (ELISA). In addition, yeast oenological traits were investigated: qualitative and technological traits were assessed on appropriate laboratory media, while the main products of microfermentation (sugars, ethanol, glycerol, acetic acid) were evaluated by Fourier transform infrared spectroscopy (FTIR). The results showed OTA reduction by 36-42% in cultures containing 100 g L⁻¹ ethanol incubated at pH 3.5 and 37 °C. OTA removal was affected by contact time, pH and ethanol content, as it was increased at low pH and by 100 g L⁻¹ ethanol. Moreover, the phenomenon was reversible, as OTA was lowest after 4 days, then it was partially released in the medium. © 2012 Society of Chemical Industry.

Influencing factors on bread-derived exposure to ochratoxin A: type, origin and composition.

PubMed

Duarte, S C; Bento, J; Pena, A; Lino, C M; Delerue-Matos, C; Oliveira, M B P P; Alves, M R; Pereira, J A

2010-01-01

The nearly ubiquitous consumption of cereals all over the world renders them an important position in international nutrition, but concurrently allocates exposure to possible contained contaminants. Mycotoxins are natural food contaminants, difficult to predict, evade, and reduce, so it is important to establish the real contribution of each contaminated food product, with the aim to evaluate mycotoxin exposure. This was the key objective of this survey and analysis for ochratoxin A content on 274 samples of commercialized bread in the Portuguese market, during the winter 2007. Different bread products were analyzed through an HPLC-FD method, including traditional types, novel segments, and different grain based bread products. A wide-ranging low level contamination was observed in all regions and types of bread products analyzed, especially in the Porto and Coimbra regions, and in the maize and whole-grain or fiber-enriched bread. Nevertheless, the exposure through contaminated wheat bread continues to be the most significant, given its high consumption and dominance in relation to the other types of bread. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

Ochratoxin A and human health risk: a review of the evidence.

PubMed

Bui-Klimke, Travis R; Wu, Felicia

2015-01-01

Ochratoxin A (OTA) is a mycotoxin produced by several fungal species including Aspergillus ochraceus, A. carbonarius, A. niger, and Penicillium verrucosum. OTA causes nephrotoxicity and renal tumors in a variety of animal species; however, human health effects are less well-characterized. Various studies have linked OTA exposure with the human diseases Balkan endemic nephropathy (BEN) and chronic interstitial nephropathy (CIN), as well as other renal diseases. This study reviews the epidemiological literature on OTA exposure and adverse health effects in different populations worldwide, and assesses the potential human health risks of OTA exposure. Epidemiological studies identified in a systematic review were used to calculate unadjusted odds ratios for OTA associated with various health endpoints. With one exception, there appears to be no statistically significant evidence for human health risks associated with OTA exposure. One Egyptian study showed a significantly higher risk of nephritic syndrome in those with very high urinary OTA levels compared with relatively unexposed individuals; however, other potential risk factors were not controlled for in the study. Larger cohort or case-control studies are needed in the future to better establish potential OTA-related human health effects, and further duplicate-diet studies are needed to validate biomarkers of OTA exposure in humans.

Ochratoxin A induced premature senescence in human renal proximal tubular cells.

PubMed

Yang, Xuan; Liu, Sheng; Huang, Chuchu; Wang, Haomiao; Luo, Yunbo; Xu, Wentao; Huang, Kunlun

2017-05-01

Ochratoxin A (OTA) has many nephrotoxic effects and is a promising compound for the study of nephrotoxicity. Human renal proximal tubular cells (HKC) are an important model for the study of renal reabsorption, renal physiology and pathology. Since the induction of OTA in renal senescence is largely unknown, whether OTA can induce renal senescence, especially at a sublethal dose, and the mechanism of OTA toxicity remain unclear. In our study, a sublethal dose of OTA led to an enhanced senescent phenotype, β-galactosidase staining and senescence associated secretory phenotype (SASP). Cell cycle arrest and cell shape alternations also confirmed senescence. In addition, telomere analysis by RT-qPCR allowed us to classify OTA-induced senescence as a premature senescence. Western blot assays showed that the p53-p21 and the p16-pRB pathways and the ezrin-associated cell spreading changes were activated during the OTA-induced senescence of HKC. In conclusion, our results demonstrate that OTA promotes the senescence of HKC through the p53-p21 and p16-pRB pathways. The understanding of the mechanisms of OTA-induced senescence is critical in determining the role of OTA in cytotoxicity and its potential carcinogenicity. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

A risk assessment of dietary Ochratoxin a in the United States.

PubMed

Mitchell, Nicole J; Chen, Chen; Palumbo, Jeffrey D; Bianchini, Andreia; Cappozzo, Jack; Stratton, Jayne; Ryu, Dojin; Wu, Felicia

2017-02-01

Ochratoxin A (OTA) is a mycotoxin (fungal toxin) found in multiple foodstuffs. Because OTA has been shown to cause kidney disease in multiple animal models, several governmental bodies around the world have set maximum allowable levels of OTA in different foods and beverages. In this study, we conducted the first exposure and risk assessment study of OTA for the United States' population. A variety of commodities from grocery stores across the US were sampled for OTA over a 2-year period. OTA exposure was calculated from the OTA concentrations in foodstuffs and consumption data for different age ranges. We calculated the margin of safety (MOS) for individual age groups across all commodities of interest. Most food and beverage samples were found to have non-detectable OTA; however, some samples of dried fruits, breakfast cereals, infant cereals, and cocoa had detectable OTA. The lifetime MOS in the US population within the upper 95% of consumers of all possible commodities was >1, indicating negligible risk. In the US, OTA exposure is highest in infants and young children who consume large amounts of oat-based cereals. Even without OTA standards in the US, exposures would not be associated with significant risk of adverse effects. Copyright © 2016 Elsevier Ltd. All rights reserved.

Level of contamination with mycobiota and contents of mycotoxins from the group of trichothecenes in grain of wheat , oats, barley, rye and triticale harvested in Poland in 2006- 2008.

PubMed

Stuper-Szablewska, Kinga; Perkowski, Juliusz

2017-03-01

The risk of cereal exposure to microbial contamination is high and possible at any time, starting from the period of plant vegetation, through harvest, up to the processing, storage and transport of the final product. Contents of mycotoxins in grain are inseparably connected with the presence of fungal biomass, the presence of which may indicate the occurrence of a fungus, and indirectly also products of its metabolism. Analyses were conducted on 378 grain samples of wheat, triticale, barley, rye and oats collected from grain silos located at grain purchase stations and at mills in Poland in 2006, 2007 and 2008. The concentrations of ERG and mycotoxins from the group of trichothecenes, as well as CFU numbers were analysed. The tested cereals were characterised by similarly low concentrations of both the investigated fungal metabolites and the level of microscopic fungi. However, conducted statistical analyses showed significant variation between tested treatments. Oat and rye grain contained the highest amounts of ERG, total toxins and CFU. In turn, the lowest values of investigated parameters were found in grain of wheat and triticale. Chemometric analyses, based on the results of chemical and microbiological tests, showed slight differences between contents of analysed metabolites between the years of the study, and do not confirm the observations on the significance of the effect of weather conditions on the development of mycobiota and production of mycotoxins; however, it does pertain to treatments showing no significant infestation. Highly significant correlations between contents of trichothecenes and ERG concentration (higher than in the case of the correlation of the total toxin concentrations/log cfu/g), indicate that the level of this metabolite is inseparably connected with mycotoxin contents in grain.

Ochratoxin A in cocoa and chocolate sampled in Canada.

PubMed

Turcotte, A-M; Scott, P M

2011-06-01

In order to determine the levels of ochratoxin A (OTA) in cocoa and cocoa products available in Canada, a previously published analytical method, with minor modifications to the extraction and immunoaffinity clean-up and inclusion of an evaporation step, was initially used (Method I). To improve the low method recoveries (46-61%), 40% methanol was then included in the aqueous sodium bicarbonate extraction solvent (pH 7.8) (Method II). Clean-up was on an Ochratest™ immunoaffinity column and OTA was determined by liquid chromatography (LC) with fluorescence detection. Recoveries of OTA from spiked cocoa powder (0.5 and 5 ng g(-1)) were 75-84%; while recoveries from chocolate were 93-94%. The optimized method was sensitive (limit of quantification (LOQ) = 0.07-0.08 ng g(-1)), accurate (recovery = 75-94%) and precise (coefficient of variation (CV) < 5%). It is applicable to cocoa and chocolate. Analysis of 32 samples of cocoa powder (16 alkalized and 16 natural) for OTA showed an incidence of 100%, with concentrations ranging from 0.25 to 7.8 ng g(-1); in six samples the OTA level exceeded 2 ng g(-1), the previously considered European Union limit for cocoa. The frequency of detection of OTA in 28 chocolate samples (21 dark or baking chocolate and seven milk chocolate) was also 100% with concentrations ranging from 0.05 to 1.4 ng g(-1); one sample had a level higher than the previously considered European Union limit for chocolate (1 ng g(-1)).

Utilising an LC-MS/MS-based multi-biomarker approach to assess mycotoxin exposure in the Bangkok metropolitan area and surrounding provinces.

PubMed

Warth, Benedikt; Petchkongkaew, Awanwee; Sulyok, Michael; Krska, Rudolf

2014-01-01

Human exposures to mycotoxins through dietary intake are a major health hazard and may result in various pathophysiological effects. Although Thailand is a country at increased risk due to its climatic conditions, no comprehensive dataset is available to perform proper exposure assessment of its population with regard to mycotoxins. Therefore, this pilot study was conducted to investigate and evaluate the exposure levels of major mycotoxins (aflatoxin B₁, ochratoxin A, fumonisins, zearalenone and trichothecenes). Sixty first-morning urine samples were collected from healthy volunteers who live in the Bangkok metropolitan area and surrounding provinces (Pathumthani, Nonthaburi, Samutprakarn and Samutsakorn). Urine samples were analysed by a LC-MS/MS-based multi-biomarker method following a so-called 'dilute and shoot' approach. Results generally indicated low mycotoxin exposures in most individuals through the determination of the four biomarkers that were detected in urine samples, i.e. aflatoxin M₁, ochratoxin A (OTA), as well as the deoxynivalenol (DON) metabolites DON-3-glucuronide and DON-15-glucuronide in 10 of 60 individuals. The maximum concentrations were used to estimate the daily intake confirming that none of the individuals exceeded the tolerable daily intake (TDI) of DON (maximum 26% of TDI) or OTA (maximum 22% of TDI). However, the maximum exposure of aflatoxin B₁, estimated to be 0.91 µg (kg bw)⁻¹ day⁻¹, should raise some concerns and suggests further studies utilising a more sensitive method. Low exposure to Fusarium toxins was also confirmed by the absence of zearalenone, α-zearalanol, β-zearalanol and zearalenone-14-glucuronide as well as T-2 toxin, HT-2 toxin, nivalenol and free DON. This is the first multi-mycotoxin biomarker study performed in Southeast Asia.

pH-Signaling Transcription Factor AopacC Regulates Ochratoxin A Biosynthesis in Aspergillus ochraceus.

PubMed

Wang, Yan; Liu, Fei; Wang, Liuqing; Wang, Qi; Selvaraj, Jonathan Nimal; Zhao, Yueju; Wang, Yun; Xing, Fuguo; Liu, Yang

2018-05-02

In Aspergillus and Penicillium species, an essential pH-response transcription factor pacC is involved in growth, pathogenicity, and toxigenicity. To investigate the connection between ochratoxin A (OTA) biosynthesis and ambient pH, the AopacC in Aspergillus ochraceus was functionally characterized using a loss-of-function mutant. The mycelium growth was inhibited under pH 4.5 and 10.0, while the sporulation increased under alkaline condition. A reduction of mycelium growth and an elevation of sporulation was observed in Δ AopacC mutant. Compared to neutral condition, OTA contents were respectively reduced by 71.6 and 79.8% under acidic and alkaline conditions. The expression of AopacC increased with the elevated pH, and deleting AopacC dramatically decreased OTA production and biosynthetic genes Aopks expression. Additionally, the Δ AopacC mutant exhibited attenuated infection ability toward pear fruits. These results suggest that AopacC is an alkaline-induced regulator responsible for growth and OTA biosynthesis in A. ochraceus and this regulatory mechanism might be pH-dependent.

Rolling chain amplification based signal-enhanced electrochemical aptasensor for ultrasensitive detection of ochratoxin A.

PubMed

Huang, Lin; Wu, Jingjing; Zheng, Lei; Qian, Haisheng; Xue, Feng; Wu, Yucheng; Pan, Daodong; Adeloju, Samuel B; Chen, Wei

2013-11-19

A novel electrochemical aptasensor is described for rapid and ultrasensitive detection of ochratoxin A (OTA) based on signal enhancement with rolling circle amplification (RCA). The primer for RCA was designed to compose of a two-part sequence, one part of the aptamer sequence directed against OTA while the other part was complementary to the capture probe on the electrode surface. In the presence of target OTA, the primer, originally hybridized with the RCA padlock, is replaced to combine with OTA. This induces the inhibition of RCA and decreases the OTA sensing signal obtained with the electrochemical aptasensor. Under the optimized conditions, ultrasensitive detection of OTA was achieved with a limit of detection (LOD) of 0.065 ppt (pg/mL), which is much lower than previously reported. The electrochemical aptasensor was also successfully applied to the determination of OTA in wine samples. This ultrasensitive electrochemical aptasensor is of great practical importance in food safety and could be widely extended to the detection of other toxins by replacing the sequence of the recognition aptamer.

Ultrasensitive one-step rapid detection of ochratoxin A by the folding-based electrochemical aptasensor.

PubMed

Wu, Jingjing; Chu, Huaqin; Mei, Zhanlong; Deng, Yi; Xue, Feng; Zheng, Lei; Chen, Wei

2012-11-13

A one-step electrochemical aptasensor using the thiol- and methylene blue- (MB-) dual-labeled aptamer modified gold electrode for determination of ochratoxin A (OTA) was presented in this research. The aptamer against OTA was covalently immobilized on the surface of the electrode by the self-assembly effect and used as recognition probes for OTA detection by the binding induced folding of the aptamer. Under the optimal conditions, the developed electrochemical aptasensor demonstrated a wide linear range from 0.1 pg mL(-1) to 1000 pg mL(-1) with the limit of detection (LOD) of 0.095 pg mL(-1), which was an extraordinary sensitivity compared with other common methods for OTA detection. Moreover, as a practical application, this proposed electrochemical aptasensor was used to monitor the OTA level in red wine samples without any special pretreatment and with satisfactory results obtained. Study results showed that this electrochemical aptasensor could be a potential useful platform for on-site OTA measurement in real complex samples. Copyright © 2012 Elsevier B.V. All rights reserved.

Acute toxicity of ochratoxin-A in marine water-reared sea bass (Dicentrarchus labrax L.).

PubMed

El-Sayed, Yasser Said; Khalil, Riad Hassan; Saad, Talaat Talaat

2009-05-01

The toxic effects of ochratoxin-A (OTA) on sea bass, Dicentrarchus labrax L., have not been previously documented. A flow-through bioassay test system was conducted in two series and a total 180 of adult marine-reared sea bass was used to estimate the acute oral 96 h median lethal concentration (LC(50)) value and behavioral changes of OTA. The data obtained were statistically evaluated using Finney's Probit Analysis Method developed by EPA. The 96 h LC(50) value for adult D. labrax was found to be 277 microg kg(-1)bwt with 95% confidence limits of 244-311 microg kg(-1)bwt. This value was calculated to be 285 microg kg(-1) bwt with Behrens-Karber's method. The two methods were relatively comparable. The acute dietary 96 h LC(50) of OTA is 9.23 mg kg(-1) diet. Additionally, the behavioral changes of sea bass were primarily observed as nervous and respiratory manifestations. We concluded that sea bass is a species highly sensitive to OTA making them a useful experimental model for aquatic mycotoxigenic problems.

Luminescence resonance energy transfer (LRET) aptasensor for ochratoxin A detection using upconversion nanoparticles

NASA Astrophysics Data System (ADS)

Jo, Eun-Jung; Byun, Ju-Young; Mun, Hyoyoung; Kim, Min-Gon

2017-07-01

We report an aptasensor for homogeneous ochratoxin A (OTA) detection based on luminescence resonance energy transfer (LRET). This system uses upconversion nanoparticles (UCNPs), such as NaYF4:Yb3+, Er 3+, as the donor. The aptamer includes the optimum-length linker (5-mer-length DNA) and OTA-specific aptamer sequences. Black hole quencher 1 (BHQ1), as the acceptor, was modified at the 3' end of the aptamer sequence. BHQ1 plays as a quencher in LRET aptasensor and shows absorption at 543 nm, which overlaps with well the emission of the UCNPs. When OTA is added, the BHQ1-labeled OTA aptamer was folded due to the formation of the G-quadruplex-OTA complex, which induced the BHQ1 close to the UCNPs. Consequently, resonance energy transfer between UCNPs (donor) and BHQ1 (acceptor) enables quenching of upconversion luminescence signals under laser irradiation of 980 nm. Our results showed that the LRET-based aptasensor allows specific OTA analysis with a limit of detection of 0.03 ng/mL. These results demonstrated that the OTA in diverse foods can be detected specifically and sensitively in a homogeneous manner.

Impedimetric Label-Free Immunosensor on Disposable Modified Screen-Printed Electrodes for Ochratoxin A

PubMed Central

Malvano, Francesca; Albanese, Donatella; Crescitelli, Alessio; Pilloton, Roberto; Esposito, Emanuela

2016-01-01

An impedimetric label-free immunosensor on disposable screen-printed carbon electrodes (SPCE) for quantitative determination of Ochratoxin A (OTA) has been developed. After modification of the SPCE surface with gold nanoparticles (AuNPs), the anti-OTA was immobilized on the working electrode through a cysteamine layer. After each coating step, the modified surfaces were characterized by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). The capacitance was chosen as the best parameter that describes the reproducible change in electrical properties of the electrode surface at different OTA concentrations and it was used to investigate the analytical parameters of the developed immunosensor. Under optimized conditions, the immunosensor showed a linear relationship between 0.3 and 20 ng/mL with a low detection limit of 0.25 ng/mL, making it suitable to control OTA content in many common food products. Lastly, the immunosensor was used to measure OTA in red wine samples and the results were compared with those registered with a competitive ELISA kit. The immunosensor was sensitive to OTA lower than 2 μg/kg, which represents the lower acceptable limit of OTA established by European legislation for common food products. PMID:27376339

Impedimetric Label-Free Immunosensor on Disposable Modified Screen-Printed Electrodes for Ochratoxin A.

PubMed

Malvano, Francesca; Albanese, Donatella; Crescitelli, Alessio; Pilloton, Roberto; Esposito, Emanuela

2016-06-30

An impedimetric label-free immunosensor on disposable screen-printed carbon electrodes (SPCE) for quantitative determination of Ochratoxin A (OTA) has been developed. After modification of the SPCE surface with gold nanoparticles (AuNPs), the anti-OTA was immobilized on the working electrode through a cysteamine layer. After each coating step, the modified surfaces were characterized by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). The capacitance was chosen as the best parameter that describes the reproducible change in electrical properties of the electrode surface at different OTA concentrations and it was used to investigate the analytical parameters of the developed immunosensor. Under optimized conditions, the immunosensor showed a linear relationship between 0.3 and 20 ng/mL with a low detection limit of 0.25 ng/mL, making it suitable to control OTA content in many common food products. Lastly, the immunosensor was used to measure OTA in red wine samples and the results were compared with those registered with a competitive ELISA kit. The immunosensor was sensitive to OTA lower than 2 μg/kg, which represents the lower acceptable limit of OTA established by European legislation for common food products.

Determination of ochratoxin A in traditional Chinese medicinal plants by HPLC-FLD.

PubMed

Yang, Lei; Wang, Linan; Pan, Jianyu; Xiang, Lan; Yang, Meihua; Logrieco, Antonio F

2010-07-01

Traditional Chinese medicinal plants (TCMPs), commonly used as spices, additives or foods, are also widely used in China to prevent and cure human disease. Due to their provenance, TCMPs may be contaminated by various fungal species, including ochratoxigenic fungi, during growth, collection, transportation and, especially, storage. A reliable method for the determination of ochratoxin A (OTA) in TCMPs of different origins was developed to monitor OTA levels in China. Analyses involved the extraction of OTA by methanol/water, immunoaffinity column (IAC) clean-up and HPLC quantification with fluorescence detection (FLD). Positive results were further confirmed by LC-ESI-MS/MS. The limit of detection (LOD) was 0.3 microg kg(-1), based on a signal-to-noise ratio of 3 : 1. Among the total of 57 TCMPs samples collected from six different areas, 31 were visibly moldy due to inappropriate storage; 26 sample, purchased from local TCMPs markets, were not visibly moldy. The results showed that 23 of the visibly moldy samples and two of the not visibly moldy were contaminated with OTA at levels ranging 1.2-158.7 and 2.5-5.6 microg kg(-1), respectively. This is the first report of the natural occurrence of OTA in TCMPs.

Ochratoxin A reduction in meat sausages using processing methods practiced in households.

PubMed

Pleadin, Jelka; Perši, Nina; Kovačević, Dragan; Vulić, Ana; Frece, Jadranka; Markov, Ksenija

2014-01-01

The aim of this study was to investigate the possibilities of ochratoxin A (OTA) reduction in home-made meat products. Meat sausages (n = 50) produced from raw materials coming from pigs exposed to OTA-contaminated feed, were subject to common heat processes practiced in households (cooking, frying and baking). Concentrations of OTA in pre- and post-processed products were quantified using a validated immunoassay method, enzyme-linked immunosorbent assay, and confirmed using a high-performance liquid chromatography with fluorescence detection. In line with the differences in recipes used and the degree of OTA accumulation in raw materials, OTA concentrations established in Mediterranean and roast sausages were lower than those found in liver and blood sausages. Baking of contaminated sausages at the temperatures of 190-220°C (for 60 min) resulted in significant reduction of OTA levels (75.8%), while 30-min cooking (at 100°C) and frying (at 170°C) proved to be significantly less effective (e.g. yielding OTA reductions of 7.4% and 12.6%, respectively). The results pointed out that despite high OTA stability, heat processes are capable of reducing its concentration in home-made meat products, depending on the processing modality used.

Ochratoxin A and its metabolites in urines of German adults-An assessment of variables in biomarker analysis.

PubMed

Ali, Nurshad; Muñoz, Katherine; Degen, Gisela H

2017-06-05

Ochratoxin A (OTA), a mycotoxin known for its nephrotoxic and carcinogenic properties, is a worldwide occurring contaminant in a variety of food commodities. Biomonitoring (i.e. analysis in biological fluids) can serve to assess human internal exposure from all consumed foods and beverages. We now determined the concentration of OTA and its metabolite ochratoxin alpha (OTα) in plasma and in urine of two male volunteers with different food habits, in order to assess intra-individual temporal fluctuations and inter-individual differences in their biomarker levels. Moreover, the urinary levels of both OTA and OTα were analyzed in a cohort of German adults (23 males, 27 females) on their regular diet. All samples were subjected to an enzymatic hydrolysis of biomarker conjugates prior to clean-up by liquid-liquid extraction and HPLC-FD analysis. The profile in the first individual showed small fluctuations over time: mean levels in plasma were 0.42 and 0.45ng/mL for OTA and OTα, respectively, and in urine means of 0.06ng/mL for both analytes. The other individual had mean levels of 1.64 and 0.20ng/mL for OTA and OTα in plasma, and 0.24 and 2.22ng/mL for these analytes in urine. It is concluded that inter-individual differences in biomarker levels reflect dissimilar dietary exposure and/or disposition of ingested mycotoxin, with an apparently more efficient detoxification of OTA to OTα in the second individual. In the German cohort (n=50), analytes were detected in 100% (OTA: range 0.02-1.82ng/mL mean level 0.21±0.31ng/mL) and 78% (OTα: range 0.01-14.25ng/mL, mean level 1.33±2.63ng/mL) of all urines. Parameters such as gender, age and body mass index did not show a significant association with urine biomarker levels. This study indicates frequent exposure to OTA among German adults. The new results are discussed in the context of biomarker data from other countries and some methodological issues. Copyright © 2017 Elsevier B.V. All rights reserved.

Formation of (4R)- and (4S)-4-hydroxyochratoxin A from ochratoxin A by liver microsomes from various species.

PubMed Central

Størmer, F C; Hansen, C E; Pedersen, J I; Hvistendahl, G; Aasen, A J

1981-01-01

Two metabolic products were formed from ochratoxin A by human, pig, and rat liver microsomal fractions in the presence of reduced nicotinamide adenine dinucleotide phosphate. They were isolated from the incubation mixture in the presence of pig liver microsomes by extraction, thin-layer chromatography, and high-pressure liquid chromatography Their structures are suggested to be (4R)- and (4S)-4-hydroxyochratoxin A on the basis of mass and nuclear magnetic resonance spectroscopy. Km and the maximum velocity for the formation of the two metabolites by human, pig, and rat microsomes were determined. Their formation was inhibited by carbon monoxide and metyrapone. The results indicate that the microsomal hydroxylation system is a cytochrome P-450 and that different species are involved in the formation of the two epimeric forms of 4-hydroxyochratoxin A. PMID:7316512

Analysis of cocoa products for ochratoxin A and aflatoxins.

PubMed

Turcotte, Anne-Marie; Scott, Peter M; Tague, Brett

2013-08-01

Eighty-five samples of cocoa products sampled in Canada were analysed for ochratoxin A (OTA) and aflatoxins in 2011-2012. Inclusion of the aflatoxins in this survey required additional method development. Chocolate was extracted with methanol-water plus NaCl, while for cocoa two successive extractions with methanol and methanol-water were made. Extracts were cleaned on an AflaOchra immunoaffinity column (IAC). Determination was by reversed phase high performance liquid chromatography (HPLC). Detection of the aflatoxins was with a post-column photochemical reactor and of OTA by fluorescence detection. Mean limits of quantification (LOQ) of chocolate and cocoa powders were 0.16 ng/g (OTA) and 0.07 ng/g (aflatoxin B1), respectively. Survey results showed that the incidences of OTA above the LOQ in natural cocoa were 15/15 (mean 1.17 ng/g), 20/21 for alkalized cocoa (mean 1.06 ng/g), 9/9 for baking chocolate (mean 0.49 ng/g), 20/20 for dark chocolate (mean 0.39 ng/g), 7/10 for milk chocolate (mean 0.19 ng/g), 5/5 for cocoa liquor (mean 0.43 ng/g), and 0/5 for cocoa butter. These results confirm our previous work with OTA. In the same samples, incidences of aflatoxin B1 above the LOQ were 14/15 for natural cocoa (mean 0.86 ng/g), 20/21 for alkalized cocoa (mean 0.37 ng/g), 7/9 for baking chocolate (mean 0.22 ng/g), 16/20 for dark chocolate (mean 0.19 ng/g), 7/10 for milk chocolate (mean 0.09 ng/g), 4/5 for cocoa liquor (mean 0.43 ng/g), and 0/5 for cocoa butter. Both aflatoxins and OTA were confirmed by HPLC-MS/MS when OTA or aflatoxin levels found were above 2 ng/g in cocoa.

Exposure of infants to ochratoxin A with breast milk.

PubMed

Muñoz, K; Blaszkewicz, M; Campos, V; Vega, M; Degen, G H

2014-03-01

The nephrotoxic and carcinogenic mycotoxin ochratoxin A (OTA) is a worldwide contaminant in food commodities and also found frequently in human biological fluids. Dietary contaminants ingested by nursing mothers can appear in breast milk. But the rate of lactational transfer of OTA has not been investigated so far at various stages of breastfeeding. Therefore, and to investigate OTA exposure of Chilean infants, we conducted a longitudinally designed study in mother-child pairs (n = 21) with parallel collection of maternal blood, milk and of infant urine samples over a period of up to 6 months. Validated analytical methods were applied to determine OTA concentrations in all biological samples (n = 134). OTA was detected in almost all maternal blood plasma, at concentrations ranging between 72 and 639 ng/L. The OTA concentrations in breast milk were on average one quarter of those measured in plasma (M/P ratio 0.25). Interestingly, a higher fraction of circulating OTA was excreted in colostrum (M/P 0.4) than with mature milk (M/P ≤ 0.2). Infants exposure was calculated as daily intake from our new data for OTA levels in breast milk, and taking into account milk consumption and body weight as additional variables: Chilean infants have an average intake of 12.7 ± 9.1 ng/kg bw during the first 6 days after delivery while intake with mature milk results in average values close to 5.0 ng/kg bw/day. Their OTA exposure is discussed in the context of tolerable intake values suggested by different scientific bodies. Moreover, the study design enabled a comparison of OTA intake and infant urine concentrations over the breastfeeding period. The statistical analysis of n = 27 paired values showed a good correlation (r = 0.57) for this type of studies and thereby confirms that urinary OTA analysis in infants is a valid biomarker of exposure.

A competitive aptamer chemiluminescence assay for ochratoxin A using a single silica photonic crystal microsphere.

PubMed

Shen, Peng; Li, Wei; Ding, Zhi; Deng, Yang; Liu, Yan; Zhu, Xuerui; Cai, Tingting; Li, Jianlin; Zheng, Tiesong

2018-08-01

We designed a competitive aptamer chemiluminescence assay for ochratoxin A (OTA) on the surface of a single silica photonic crystal microsphere (SPCM) in cereal samples. The structural color of SPCMs is used to recognize and trace the microspheres during process of detection. Anti-aptamer was immobilized on the surface of SPCM. OTA and anti-aptamer competed to bind to aptamer when OTA and its aptamer (labeled by biotin at 5'end) were added in the system. The chemiluminescence signal was developed by the horseradish peroxidase (HRP), luminol and H 2 O 2 . The molecules on the single SPCM can produce enough chemiluminescence signal intensity for quantitative detection for OTA. The linear detection range for OTA was from 1 pg/mL to 1 ng/mL and recovery rates were 89%-95%, 81%-92% and 94%-105% in rice, wheat and corn, respectively. The results showed that the developed method for OTA using a single SPCM has a great application potential in cereal samples. Copyright © 2018 Elsevier Inc. All rights reserved.

A fluorescent aptasensor based on a DNA pyramid nanostructure for ultrasensitive detection of ochratoxin A.

PubMed

Nameghi, Morteza Alinezhad; Danesh, Noor Mohammad; Ramezani, Mohammad; Hassani, Faezeh Vahdati; Abnous, Khalil; Taghdisi, Seyed Mohammad

2016-08-01

Analytical techniques for detection of ochratoxin A (OTA) in food products and blood serum are of great significance. In this study, a fluorescent aptasensor was developed for sensitive and specific detection of OTA, based on a DNA pyramid nanostructure (DPN) and PicoGreen (PG) dye. The designed aptasensor inherits characteristics of DPN, such as high stability and capacity for PG loading. PG, as a fluorescent dye, could bind to double-stranded DNA (dsDNA). In the absence of OTA, the pyramid structure of DPN remains intact, leading to a very strong fluorescence emission. Because of higher affinity of aptamer for its target relative to its complementary strand, upon addition of target, the pyramid structure of DPN is disassembled, leading to a weak fluorescence emission. The presented aptasensor showed high specificity toward OTA with a limit of detection (LOD) as low as 0.135 nM. Besides, the designed sensing strategy was successfully utilized to recognize OTA in serum and grape juice with LODs of 0.184 and 0.149 nM, respectively.

Feeding a diet contaminated with ochratoxin A for chickens at the maximum level recommended by the EU for poultry feeds (0.1 mg/kg). 1. Effects on growth and slaughter performance, haematological and serum traits.

PubMed

Pozzo, L; Salamano, G; Mellia, E; Gennero, M S; Doglione, L; Cavallarin, L; Tarantola, M; Forneris, G; Schiavone, A

2013-05-01

The European Commission Recommendation 2006/576/EC, suggests that the maximum level of Ochratoxin A (OTA) in poultry feeds should be set at 0.1 mg OTA/kg. Thirty-six one-day-old male Hubburd broiler chickens were divided into two groups, a Control (basal diet) and an Ochratoxin A (basal diet + 0.1 mg OTA/kg) group. The growth and slaughter performance traits were recorded. The liver, spleen, bursa of Fabricius and thymus weights were measured. The erythrocyte and leukocyte numbers were assayed in blood samples, and the heterophils to lymphocytes (H/L) ratio was determined. Alpha-1-acid glycoprotein (AGP), lysozyme, the total protein and the electrophoretic pattern were evaluated in serum samples. Liver enzymes (alanino aminotransferase, ALT and aspartate aminotransferase, AST) and kidney function parameters (uric acid and creatinine) were quantified. The results revealed that feeding a 0.1 mg OTA/kg contaminated diet to chicks caused a decrease in the absolute thymus weight (p < 0.05) and a lower total protein (p < 0.01), albumin (p < 0.01), alpha (p < 0.05), beta (p = 0.001) and gamma (p = 0.001) globulins serum concentration in the Ochratoxin A group. Moreover, the albumin-to-globulin (A/G) ratio of the OTA-treated animals resulted to be higher (p < 0.05). Feeding broiler chickens, a diet contaminated with the maximum level admitted by the European Commission Recommendation (0.1 mg OTA/kg), did not affect the animal performance, slaughter traits, organ weights, haematological parameters, liver enzyme or renal function parameters concentrations but had an overall immunosuppressant effect, with reduction in the thymus weight and of the total serum protein, albumin, alpha, beta and gamma globulins concentration. Journal of Animal Physiology and Animal Nutrition © 2013 Blackwell Verlag GmbH.

Indole-diterpenes and ergot alkaloids in Cynodon dactylon (Bermuda grass) infected with Claviceps cynodontis from an outbreak of tremors in cattle.

PubMed

Uhlig, Silvio; Botha, Christo J; Vrålstad, Trude; Rolén, Elin; Miles, Christopher O

2009-12-09

Tremorgenic syndromes in mammals are commonly associated with indole-diterpenoid alkaloids of fungal origin. Cattle are sometimes affected by tremors (also called "staggers") when they graze on toxic grass pastures, and Bermuda grass ( Cynodon dactylon , kweek) has been known to be associated with tremors for several decades. This study reports the identification of paspalitrems and paspaline-like indole-diterpenes in the seedheads of Claviceps cynodontis -infected Bermuda grass collected from a pasture that had caused a staggers syndrome in cattle in South Africa and thereby links the condition to specific mycotoxins. The highest concentration (about 150 mg/kg) was found for paspalitrem B. Ergonovine and ergine (lysergic acid amide), together with their C-8 epimers, were found to co-occur with the indole-diterpenes at concentrations of about 10 microg/kg. The indole-diterpene profile of the extract from the ergotized Bermuda grass was similar to that of Claviceps paspali sclerotia. However, the C. paspali sclerotia contained in addition agroclavine and elymoclavine. This is the first study linking tremors associated with grazing of Bermuda grass to specific tremorgenic indole-diterpenoid mycotoxins.

Detection of ochratoxin A (OTA) in coffee using chemiluminescence resonance energy transfer (CRET) aptasensor.

PubMed

Jo, Eun-Jung; Mun, Hyoyoung; Kim, Su-Ji; Shim, Won-Bo; Kim, Min-Gon

2016-03-01

We report a chemiluminescence resonance energy transfer (CRET) aptasensor for the detection of ochratoxin A (OTA) in roasted coffee beans. The aptamer sequences used in this study are 5'-DNAzyme-Linker-OTA aptamer-3'-dabcyl. Dabcyl at the end of the OTA aptamer region plays as a quencher in CRET aptasensor. When hemin and OTA are added, the dabcyl-labeled OTA aptamer approaches to the G-quadruplex-hemin complex by formation of the G-quadruplex-OTA complex. The G-quadruplex-hemin complexes possess horseradish peroxidase (HRP)-like activity, and therefore, the HRP-mimicking DNAzyme (HRPzyme) catalyzes peroxidation in the presence of luminol and H2O2. Resonance energy transfer between luminol (donor) and dabcyl (acceptor) enables quenching of chemiluminescence signals. The signal decreases with increasing the concentration of OTA within the range of 0.1-100ngmL(-1) (limit of detection 0.22ngmL(-1)), and the level of recovery of the respective 1ngmL(-1) and 10ngmL(-1) spiked coffee samples was 71.5% and 93.3%. These results demonstrated the potential of the proposed method for OTA analysis in diverse foods. Copyright © 2015 Elsevier Ltd. All rights reserved.

Determination of ochratoxin A and T-2 toxin in alcoholic beverages by hollow fiber liquid phase microextraction and ultra high-pressure liquid chromatography coupled to tandem mass spectrometry.

PubMed

Romero-González, R; Frenich, A Garrido; Vidal, J L Martínez; Aguilera-Luiz, M M

2010-06-30

A new method for the determination of ochratoxin A and T-2 toxin in alcoholic beverages (wine and beer) by hollow fiber liquid microextraction was optimized. The extraction step was followed by ultra high-pressure liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS). The extraction procedure was based on the extraction of mycotoxins from the sample to the organic solvent (1-octanol) immobilized in the fiber, and afterwards, they were desorbed in a mixture of acetonitrile/water (80:20, v/v) at pH 7 prior to chromatographic determination. Different variables affecting the extraction process such as organic solvent, salt content, extraction time and desorption solution were studied. The developed method was validated in wine and beer, using white wine and alcoholic beer as representative matrices for both types of samples. Relative recoveries higher than 70% were obtained for the selected mycotoxins. Good linearity (R(2)>0.993) was obtained and quantification limits (0.02-0.09 microg L(-1)) below European regulatory levels were achieved. Repeatability, expressed as relative standard deviation, was always lower than 12%, whereas interday precision was lower than 21%. The proposed method was applied to the analysis of several types of wines and beers and ochratoxin A was detected in a rosé wine at 1.1 microg L(-1). Copyright 2010 Elsevier B.V. All rights reserved.

Decontamination of poultry feed from ochratoxin A by UV and sunlight radiations.

PubMed

Ameer Sumbal, Gul; Hussain Shar, Zahid; Hussain Sherazi, Syed Tufail; Sirajuddin; Nizamani, Shafi Muhammad; Mahesar, Safaraz Ahmed

2016-06-01

Mycotoxin-contaminated feed is very dangerous for the growth and even life of poultry. The objective of the current study was to investigate the efficacy of ultra-violet irradiation for decontamination of ochratoxin A (OTA) in spiked and naturally contaminated poultry feed samples. Spiked and naturally contaminated feed samples were irradiated with ultra-violet light (UV) at distance of 25 cm over the feed samples. In vitro, the effect of UV intensity (0.1 mW cm(-2) at 254 nm UV-C) on different types of poultry feeds contaminated with OTA was evaluated. The same samples were also irradiated with sunlight and analysed for OTA by an indirect enzyme linked immunosorbent assay method. Poultry feed samples containing 500 µg kg(-1) were 100% decontaminated in 180 min with UV radiation while OTA was decreased to 70-95 µg kg(-1) using the same poultry feed samples after 8 h sunlight irradiation. Therefore, UV light was found to be more effective. Only 1 h of UV irradiation was found to be sufficient to bring the OTA level to the maximum regulatory limit suggested for poultry feeds (100 µg kg(-1) ), while 8 h were needed to obtain this level using sunlight radiations. The proposed approach is a viable option to reduce the level of OTA in contaminated poultry feeds. © 2015 Society of Chemical Industry. © 2015 Society of Chemical Industry.

Novel highly-performing immunosensor-based strategy for ochratoxin A detection in wine samples.

PubMed

Prieto-Simón, Beatriz; Campàs, Mònica; Marty, Jean-Louis; Noguer, Thierry

2008-02-28

The increasing concern about ochratoxin A (OTA) contamination of different food and feedstuffs demands high-performing detection techniques for quality assessment. Two indirect competitive enzyme-linked immunosorbent assay (ELISA) strategies were investigated for the development of OTA electrochemical immunosensors based on different OTA immobilisation procedures. Immunosensors based on avidin/biotin-OTA showed enhanced performance characteristics compared to those based on the adsorption of bovine serum albumin (BSA)-OTA conjugate. Performance of polyclonal (PAb) and monoclonal (MAb) antibodies against OTA was compared, showing at least one-order of magnitude lower IC(50) values when working with MAb. Alkaline phosphatase (ALP)- and horseradish peroxidase (HRP)-labelled secondary antibodies were evaluated. Both conjugates led to similar results when working with OTA standard solutions in buffer. However, whereas electroactive interferences present in spiked wine samples did not affect HRP-labelled immunosensors (4% slope deviation), they were likely oxidised at 0.225 V versus Ag/AgCl, the working potential for ALP-labelled immunosensors (25% slope deviation). Considering 80% of antibody binding as the limit of detection, values of 0.7 and 0.3 ng/mL for HRP- and ALP-labelled immunosensors respectively, validate these immunosensors as useful screening tools to assess OTA levels in wine.

Occurrence of ochratoxin A in spices commercialized in Abidjan (Côte d'Ivoire).

PubMed

Manda, Pierre; Adanou, Ketty Michele; Ardjouma, Dembelé; Adepo, Aholia Jean Baptiste; Dano, Djédjé Sébastien

2016-08-01

Ochratoxin A (OTA) is a mycotoxin produced mostly by several species of Aspergillus and Penicillium. OTA is nephrotoxic in all animal species in which it has been tested and is cancerogenic in rodents. It is associated with Balkan endemic nephropathy. It is naturally present in many crop products such as cereals (barley, wheat, maize) and dried fruits, spices, coffee, wine, olives, and cocoa. The aim of this study was to assess the contamination of three Ivoirian spices with OTA (ginger, chili, and pepper) widely consumed by the population. A total of 90 spice samples (ginger: n = 30; chili: n = 30; pepper n = 30) was taken from various sales outlets of Abidjan. OTA was quantified using an HPLC apparatus coupled with a fluorimetric detector. The chili and ginger samples were contaminated with OTA at a mean concentration of 57.48 ± 174 and 0.12 ± 0.15 μg/kg, respectively. No contamination of the pepper samples was detected. Eight (26.67 %) of the chili samples exceeded the maximum limit of 15 μg/kg established by European regulation. These results should serve as an alert on the risk to the consumer population of these products that are highly contaminated with OTA.

Cellular Development Associated with Induced Mycotoxin Synthesis in the Filamentous Fungus Fusarium graminearum

PubMed Central

Menke, Jon; Weber, Jakob; Broz, Karen; Kistler, H. Corby

2013-01-01

Several species of the filamentous fungus Fusarium colonize plants and produce toxic small molecules that contaminate agricultural products, rendering them unsuitable for consumption. Among the most destructive of these species is F. graminearum, which causes disease in wheat and barley and often infests the grain with harmful trichothecene mycotoxins. Synthesis of these secondary metabolites is induced during plant infection or in culture in response to chemical signals. Our results show that trichothecene biosynthesis involves a complex developmental process that includes dynamic changes in cell morphology and the biogenesis of novel subcellular structures. Two cytochrome P-450 oxygenases (Tri4p and Tri1p) involved in early and late steps in trichothecene biosynthesis were tagged with fluorescent proteins and shown to co-localize to vesicles we provisionally call “toxisomes.” Toxisomes, the inferred site of trichothecene biosynthesis, dynamically interact with motile vesicles containing a predicted major facilitator superfamily protein (Tri12p) previously implicated in trichothecene export and tolerance. The immediate isoprenoid precursor of trichothecenes is the primary metabolite farnesyl pyrophosphate. Changes occur in the cellular localization of the isoprenoid biosynthetic enzyme HMG CoA reductase when cultures non-induced for trichothecene biosynthesis are transferred to trichothecene biosynthesis inducing medium. Initially localized in the cellular endomembrane system, HMG CoA reductase, upon induction of trichothecene biosynthesis, increasingly is targeted to toxisomes. Metabolic pathways of primary and secondary metabolism thus may be coordinated and co-localized under conditions when trichothecene biosynthesis occurs. PMID:23667578

Managing ochratoxin A risk in the grape-wine food chain.

PubMed

Visconti, Angelo; Perrone, Giancarlo; Cozzi, Giuseppe; Solfrizzo, Michele

2008-02-01

The main source of ochratoxin A (OTA) in the wine food chain is the infection of grapes by "black aspergilli" in the field. OTA-producing black aspergilli include principally Aspergillus carbonarius, followed by A. niger and possibly A. tubingensis. They are opportunistic fungi that develop particularly on damaged berries at ripening, although they may occur and form OTA on grapes from veraison to harvest. Climatic conditions (high humidity and temperature) and geographical location are important factors favouring OTA accumulation in grape berries. The severity of aspergillus rot is influenced by excessive irrigation and rainfall prior to harvest, which causes berry splitting. In addition, berry wounds caused by insect attack provide preferential entries for black aspergilli. High OTA levels occur in grapes severely damaged by the grape moth, Lobesia botrana, particularly in Mediterranean areas. Some grape varieties display greater susceptibility to aspergillus rot due to intrinsic genetic characteristics and bunch conformation (i.e. compact>sparse). Control measures for toxigenic mycoflora in the vineyards must consider these critical control points. Proper fungicidal and insecticidal treatments can reduce OTA contamination. Nevertheless, knowledge about the fate of OTA and its distribution in wine and winery by-products is important to manage OTA risk in contaminated stock. In our wine-making experiments, only 4% of the OTA present in grapes remained in the wine--the majority is retained in pressed grape pomaces. OTA concentration remained unchanged in wine after a 1-year aging as well as in all liquid fractions collected during vinification (i.e. must, free run wine, and wine after first and second decantation). Activated carbon can reduce OTA levels in wine but negatively affects wine quality.

Natural Co-Occurrence of Mycotoxins in Foods and Feeds and Their in vitro Combined Toxicological Effects.

PubMed

Smith, Marie-Caroline; Madec, Stéphanie; Coton, Emmanuel; Hymery, Nolwenn

2016-03-26

Some foods and feeds are often contaminated by numerous mycotoxins, but most studies have focused on the occurrence and toxicology of a single mycotoxin. Regulations throughout the world do not consider the combined effects of mycotoxins. However, several surveys have reported the natural co-occurrence of mycotoxins from all over the world. Most of the published data has concerned the major mycotoxins aflatoxins (AFs), ochratoxin A (OTA), zearalenone (ZEA), fumonisins (FUM) and trichothecenes (TCTs), especially deoxynivalenol (DON). Concerning cereals and derived cereal product samples, among the 127 mycotoxin combinations described in the literature, AFs+FUM, DON+ZEA, AFs+OTA, and FUM+ZEA are the most observed. However, only a few studies specified the number of co-occurring mycotoxins with the percentage of the co-contaminated samples, as well as the main combinations found. Studies of mycotoxin combination toxicity showed antagonist, additive or synergic effects depending on the tested species, cell model or mixture, and were not necessarily time- or dose-dependent. This review summarizes the findings on mycotoxins and their co-occurrence in various foods and feeds from all over the world as well as in vitro experimental data on their combined toxicity.

Natural Co-Occurrence of Mycotoxins in Foods and Feeds and Their in vitro Combined Toxicological Effects

PubMed Central

Smith, Marie-Caroline; Madec, Stéphanie; Coton, Emmanuel; Hymery, Nolwenn

2016-01-01

Some foods and feeds are often contaminated by numerous mycotoxins, but most studies have focused on the occurrence and toxicology of a single mycotoxin. Regulations throughout the world do not consider the combined effects of mycotoxins. However, several surveys have reported the natural co-occurrence of mycotoxins from all over the world. Most of the published data has concerned the major mycotoxins aflatoxins (AFs), ochratoxin A (OTA), zearalenone (ZEA), fumonisins (FUM) and trichothecenes (TCTs), especially deoxynivalenol (DON). Concerning cereals and derived cereal product samples, among the 127 mycotoxin combinations described in the literature, AFs+FUM, DON+ZEA, AFs+OTA, and FUM+ZEA are the most observed. However, only a few studies specified the number of co-occurring mycotoxins with the percentage of the co-contaminated samples, as well as the main combinations found. Studies of mycotoxin combination toxicity showed antagonist, additive or synergic effects depending on the tested species, cell model or mixture, and were not necessarily time- or dose-dependent. This review summarizes the findings on mycotoxins and their co-occurrence in various foods and feeds from all over the world as well as in vitro experimental data on their combined toxicity. PMID:27023609

Mycotoxins co-contamination: Methodological aspects and biological relevance of combined toxicity studies.

PubMed

Alassane-Kpembi, Imourana; Schatzmayr, Gerd; Taranu, Ionelia; Marin, Daniela; Puel, Olivier; Oswald, Isabelle Paule

2017-11-02

Mycotoxins are secondary fungal metabolites produced mainly by Aspergillus, Penicillium, and Fusarium. As evidenced by large-scale surveys, humans and animals are simultaneously exposed to several mycotoxins. Simultaneous exposure could result in synergistic, additive or antagonistic effects. However, most toxicity studies addressed the effects of mycotoxins separately. We present the experimental designs and we discuss the conclusions drawn from in vitro experiments exploring toxicological interactions of mycotoxins. We report more than 80 publications related to mycotoxin interactions. The studies explored combinations involving the regulated groups of mycotoxins, especially aflatoxins, ochratoxins, fumonisins, zearalenone and trichothecenes, but also the "emerging" mycotoxins beauvericin and enniatins. Over 50 publications are based on the arithmetic model of additivity. Few studies used the factorial designs or the theoretical biology-based models of additivity. The latter approaches are gaining increased attention. These analyses allow determination of the type of interaction and, optionally, its magnitude. The type of interaction reported for mycotoxin combinations depended on several factors, in particular cell models and the tested dose ranges. However, synergy among Fusarium toxins was highlighted in several studies. This review indicates that well-addressed in vitro studies remain valuable tools for the screening of interactive potential in mycotoxin mixtures.

Chlorogenic acid and maize ear rot resistance: a dynamic study investigating Fusarium graminearum development, deoxynivalenol production, and phenolic acid accumulation.

PubMed

Atanasova-Penichon, Vessela; Pons, Sebastien; Pinson-Gadais, Laetitia; Picot, Adeline; Marchegay, Gisèle; Bonnin-Verdal, Marie-Noelle; Ducos, Christine; Barreau, Christian; Roucolle, Joel; Sehabiague, Pierre; Carolo, Pierre; Richard-Forget, Florence

2012-12-01

Fusarium graminearum is the causal agent of Gibberella ear rot and produces trichothecene mycotoxins. Basic questions remain unanswered regarding the kernel stages associated with trichothecene biosynthesis and the kernel metabolites potentially involved in the regulation of trichothecene production in planta. In a two-year field study, F. graminearum growth, trichothecene accumulation, and phenolic acid composition were monitored in developing maize kernels of a susceptible and a moderately resistant variety using quantitative polymerase chain reaction and liquid chromatography coupled with photodiode array or mass spectrometry detection. Infection started as early as the blister stage and proceeded slowly until the dough stage. Then, a peak of trichothecene accumulation occurred and infection progressed exponentially until the final harvest time. Both F. graminearum growth and trichothecene production were drastically reduced in the moderately resistant variety. We found that chlorogenic acid is more abundant in the moderately resistant variety, with levels spiking in the earliest kernel stages induced by Fusarium infection. This is the first report that precisely describes the kernel stage associated with the initiation of trichothecene production and provides in planta evidence that chlorogenic acid may play a role in maize resistance to Gibberella ear rot and trichothecene accumulation.

Effects of δ-tocotrienol on ochratoxin A-induced nephrotoxicity in rats.

PubMed

Damiano, Sara; Navas, Luigi; Lombari, Patrizia; Montagnaro, Serena; Forte, Iris M; Giordano, Antonio; Florio, Salvatore; Ciarcia, Roberto

2018-05-18

Ochratoxin A (OTA), is a natural contaminant of the food chain worldwide involved in the development of different type of cancers in animals and humans. Several studies suggested that oxidative damage might contribute to increase the cytotoxicity and carcinogenicity capabilities of OTA. The aim of this study was to evaluate the possible protective effect of δ-tocotrienol (Delta), a natural form of vitamin E, against OTA-induced nephrotoxicity. Male Sprague-Dawley rats were treated with OTA and/or Delta by gavage for 14 days. Our results shown that OTA treatment induced the increase of reactive oxigen species production correlated to a strong reduction of Glomerular Filtration Rate (GFR) and absoluted fluid reabsorption (Jv) with conseguent significant increase in blood pressure. Consistent, we noted in the kidney of rats treated with OTA, an increase in malondialdheyde and dihydroethidium production and a reduction of the activity of the catalase, superoxide dismutase, and glutathione peroxidase. Conversly, in the rat group subjected to the concomitant treatment OTA plus Delta, we observed the restored effect, compared the OTA treatment group, on blood pressure, GFR, Jv, and all activities of renal antioxidant enzymes. Finally, as far as concern the tissue damage induced by OTA and measured evaluating fibronectin protein levels, we observed that in OTA plus Delta group this effect is not restored. Our findings releval that a mechanism underlying the renal toxicity induced by OTA is the oxidative stress and provide a new rationale to use a Delta in order to protect, at least in part, against OTA-induced nephrotoxicity. © 2018 Wiley Periodicals, Inc.

Development of the custom polymeric materials specific for aflatoxin B1 and ochratoxin A for application with the ToxiQuant T1 sensor tool.

PubMed

Piletska, Elena; Karim, Kal; Coker, Raymond; Piletsky, Sergey

2010-04-16

Two polymers were computationally designed with affinity to two of the most abundant mycotoxins aflatoxin B1 (AFB1) and ochratoxin A (OTA) for application in the ToxiQuant T1 System. The principle of quantification of AFB1 and OTA using the ToxiQuant T1 instrument comprised of a fluorimetric analysis of mycotoxins adsorbed on the polymer upon exposure to UV light. High affinity of the developed resins allowed the adsorption of both toxins as discrete bands on the top of the cartridge with detection limit as low as 1ng quantity of mycotoxins. Copyright 2009 Elsevier B.V. All rights reserved.

Long term administration of low doses of mycotoxins in poultry. 2. Residues of ochratoxin A and aflatoxins in broilers and laying hens after combined administration of ochratoxin A and aflatoxin B1.

PubMed

Micco, C; Miraglia, M; Benelli, L; Onori, R; Ioppolo, A; Mantovani, A

1988-01-01

The effects of combined administration of ochratoxin A (OA) and aflatoxin B1 (AFB1) on the occurrence and the levels of residues of mycotoxins in poultry have been investigated. Male broilers and laying hens were fed from 14 days old with standard diets contaminated with 50 micrograms/kg OA and 50 micrograms/kg AFB1. Two groups of broilers and hens were withdrawn from contaminated feed at 37 and 88 days, respectively. At the time of sacrifice no significant lesions were found. Residues were compared with those found after administration of either toxin alone in former trials. Combined treatment resulted in higher content of OA in broiler livers (40 versus 5.0 micrograms/kg) and, to a lesser extent, in kidneys and skin, and of AFB1 in broiler liver and kidney (0.15 versus 0.02 microgram/kg and 0.40 versus 0.05 microgram/kg respectively). Laying hens showed smaller differences (0.20 versus 0.10 microgram/kg in liver and 0.32 versus 0.08 in kidneys). Withdrawal from treatment led to the almost complete disappearance of OA residues in broilers and in hens. These results show a synergistic effect of OA and AFB1, particularly in broilers.

Occurrence of Penicillium verrucosum, ochratoxin A, ochratoxin B and citrinin in on-farm stored winter wheat from the Canadian Great Lakes Region

PubMed Central

Miller, J. David; Schaafsma, Arthur W.

2017-01-01

The occurrence of P. verrucosum and ochratoxin A (OTA) were surveyed for 3 and 4 years, respectively. A total of 250 samples was collected from an average of 30 farms during the 2011, 2012, 2013 and 2014 winter seasons. Most storage bins surveyed were typically 11 m high round bins made of corrugated, galvanized steel, with flat-bottoms and conical roofs. Samples of clumped grain contained the most P. verrucosum (p<0.05, n = 10) followed by samples taken from the first load (n = 24, mean = 147±87 CFU/g) and last load (n = 17, mean = 101±77 CFU/g). Five grain samples (2.2%) tested positive for OTA, citrinin and OTB at concentrations of 14.7±7.9, 4.9±1.9 and 1.2±0.7 ng/g, with only three samples exceeding 5 ng/g. Grain samples positive for OTA were related to moisture resulting from either condensation or migrating moist warm air in the bin or areas where precipitation including snow entered the bin. Bins containing grain and clumps contaminated with OTA were studied in detail. A number of statistically-significant risk factors for OTA contamination were identified. These included 1) grain clumps accumulated around or directly under manhole openings, 2) debris and residue of old grain or grain clumps collected from the bin walls or left on storage floor and augers and 3) grain clumps accumulated around side doors. Even when grain enters storage below the 14.5% threshold of moisture, condensation and moisture migration occurs in hotspots in modern corrugated steel storage bins. Hot spots of OTA contamination were most often in areas affected by moisture migration due to inadequate aeration and exposure to moisture from precipitation or condensation. Further, we found that the nature of the condensation affects the nature and distribution of small and isolated areas with high incidence of toxin contamination and/or P. verrucosum prevalence in the grain bins examined. PMID:28749978

Occurrence of Penicillium verrucosum, ochratoxin A, ochratoxin B and citrinin in on-farm stored winter wheat from the Canadian Great Lakes Region.

PubMed

Limay-Rios, Victor; Miller, J David; Schaafsma, Arthur W

2017-01-01

The occurrence of P. verrucosum and ochratoxin A (OTA) were surveyed for 3 and 4 years, respectively. A total of 250 samples was collected from an average of 30 farms during the 2011, 2012, 2013 and 2014 winter seasons. Most storage bins surveyed were typically 11 m high round bins made of corrugated, galvanized steel, with flat-bottoms and conical roofs. Samples of clumped grain contained the most P. verrucosum (p<0.05, n = 10) followed by samples taken from the first load (n = 24, mean = 147±87 CFU/g) and last load (n = 17, mean = 101±77 CFU/g). Five grain samples (2.2%) tested positive for OTA, citrinin and OTB at concentrations of 14.7±7.9, 4.9±1.9 and 1.2±0.7 ng/g, with only three samples exceeding 5 ng/g. Grain samples positive for OTA were related to moisture resulting from either condensation or migrating moist warm air in the bin or areas where precipitation including snow entered the bin. Bins containing grain and clumps contaminated with OTA were studied in detail. A number of statistically-significant risk factors for OTA contamination were identified. These included 1) grain clumps accumulated around or directly under manhole openings, 2) debris and residue of old grain or grain clumps collected from the bin walls or left on storage floor and augers and 3) grain clumps accumulated around side doors. Even when grain enters storage below the 14.5% threshold of moisture, condensation and moisture migration occurs in hotspots in modern corrugated steel storage bins. Hot spots of OTA contamination were most often in areas affected by moisture migration due to inadequate aeration and exposure to moisture from precipitation or condensation. Further, we found that the nature of the condensation affects the nature and distribution of small and isolated areas with high incidence of toxin contamination and/or P. verrucosum prevalence in the grain bins examined.

Quantitative and qualitative transcriptome analysis of four industrial strains of Claviceps purpurea with respect to ergot alkaloid production.

PubMed

Majeská Čudejková, Mária; Vojta, Petr; Valík, Josef; Galuszka, Petr

2016-09-25

The fungus Claviceps purpurea is a biotrophic phytopathogen widely used in the pharmaceutical industry for its ability to produce ergot alkaloids (EAs). The fungus attacks unfertilized ovaries of grasses and forms sclerotia, which represent the only type of tissue where the synthesis of EAs occurs. The biosynthetic pathway of EAs has been extensively studied; however, little is known concerning its regulation. Here, we present the quantitative transcriptome analysis of the sclerotial and mycelial tissues providing a comprehensive view of transcriptional differences between the tissues that produce EAs and those that do not produce EAs and the pathogenic and non-pathogenic lifestyle. The results indicate metabolic changes coupled with sclerotial differentiation, which are likely needed as initiation factors for EA biosynthesis. One of the promising factors seems to be oxidative stress. Here, we focus on the identification of putative transcription factors and regulators involved in sclerotial differentiation, which might be involved in EA biosynthesis. To shed more light on the regulation of EA composition, whole transcriptome analysis of four industrial strains differing in their alkaloid spectra was performed. The results support the hypothesis proposing the composition of the amino acid pool in sclerotia to be an important factor regulating the final structure of the ergopeptines produced by Claviceps purpurea. Copyright © 2016 Elsevier B.V. All rights reserved.

Influence of Fermentation and Drying Materials on the Contamination of Cocoa Beans by Ochratoxin A

PubMed Central

Dano, Sébastien Djédjé; Manda, Pierre; Dembélé, Ardjourma; Abla, Ange Marie-Joseph Kouassi; Bibaud, Joel Henri; Gouet, Julien Zroh; Sika, Charles Bruno Ze Maria

2013-01-01

Ochratoxin A (OTA) is a mycotoxin produced mainly by species of Aspergillus and Penicillium. Contamination of food with OTA is a major consumer health hazard. In Côte d’Ivoire, preventing OTA contamination has been the subject of extensive study. The current study was conducted to evaluate the influence of fermentation and drying materials on the OTA content in cocoa. For each test, 7000 intact cocoa pods were collected, split open to remove the beans, fermented using 1 of 3 different materials, sun-dried on 1 of 3 different platform types and stored for 30 days. A total of 22 samples were collected at each stage of post-harvesting operations. The OTA content in the extracted samples was then quantified by high-performance liquid chromatography. OTA was detected in beans at all stages of post-harvesting operations at varying levels: pod-opening (0.025 ± 0.02 mg/kg), fermentation (0.275 ± 0.2 mg/kg), drying (0.569 ± 0.015 mg/kg), and storage (0.558 ± 0.04 mg/kg). No significant relationships between the detected OTA level and the materials used in the fermentation and drying of cocoa were observed. PMID:24287569

Efficacy of natamycin for control of growth and ochratoxin A production by Aspergillus carbonarius strains under different environmental conditions.

PubMed

Medina, A; Jiménez, M; Mateo, R; Magan, N

2007-12-01

To examine the efficacy of natamycin produced by Streptomyces natalensis against strains of Aspergillus carbonarius growth and ochratoxin A (OTA) production under different environmental factors on a grape juice-based medium. Detailed studies in the range 0-20 ng ml(-1) for control of growth and ochratoxin production by strains of A. carbonarius at 0.98, 0.96 and 0.94 water availabilities (a(w)) and 15-25 degrees C on a fresh red grape extract medium were examined. Inhibition of growth was depending on temperature and a(w) level. At 15 degrees C, 5-10 ng ml(-1) natamycin was effective in reducing growth almost completely. However, at 20-25 degrees C and all the three a(w) levels, growth was only slightly inhibited by 5-10 ng ml(-1) natamycin. There were strain differences with regard to inhibition of OTA production. At 15 degrees C and 0.98 a(w), 10 ng ml(-1) was required to inhibit production by >90%. However, at 0.96 and 0.94 a(w), almost complete inhibition occurred. At 20 degrees C, OTA production was only significantly inhibited by 10 ng ml(-1) natamycin at 0.94 a(w). At 0.96 and 0.98 a(w), some inhibition occurred with 5-10 ng ml(-1), but greater concentrations would be required for effective inhibition. At 25 degrees C, 5 ng ml(-1) was effective at all a(w) levels. However, at 15 degrees C and 25 degrees C and a wide range of a(w) levels, natamycin effectively controlled OTA production. Natamycin appears to be a very effective for controlling growth and OTA production by strains of A. carbonarius over a range of a(w) and temperature conditions on grape-based media. This is the first detailed study to demonstrate the impact of natamycin against A. carbonarius. This study suggests that use of natamycin at 50-100 ng ml(-1) can give complete inhibition of growth of A. carbonarius and OTA production over a range of environmental conditions. Natamycin could be an important component of a system to prevent OTA contamination of wine as well during the drying and

Detection of ochratoxin A contamination in stored wheat using near-infrared hyperspectral imaging

NASA Astrophysics Data System (ADS)

Senthilkumar, T.; Jayas, D. S.; White, N. D. G.; Fields, P. G.; Gräfenhan, T.

2017-03-01

Near-infrared (NIR) hyperspectral imaging system was used to detect five concentration levels of ochratoxin A (OTA) in contaminated wheat kernels. The wheat kernels artificially inoculated with two different OTA producing Penicillium verrucosum strains, two different non-toxigenic P. verrucosum strains, and sterile control wheat kernels were subjected to NIR hyperspectral imaging. The acquired three-dimensional data were reshaped into readable two-dimensional data. Principal Component Analysis (PCA) was applied to the two dimensional data to identify the key wavelengths which had greater significance in detecting OTA contamination in wheat. Statistical and histogram features extracted at the key wavelengths were used in the linear, quadratic and Mahalanobis statistical discriminant models to differentiate between sterile control, five concentration levels of OTA contamination in wheat kernels, and five infection levels of non-OTA producing P. verrucosum inoculated wheat kernels. The classification models differentiated sterile control samples from OTA contaminated wheat kernels and non-OTA producing P. verrucosum inoculated wheat kernels with a 100% accuracy. The classification models also differentiated between five concentration levels of OTA contaminated wheat kernels and between five infection levels of non-OTA producing P. verrucosum inoculated wheat kernels with a correct classification of more than 98%. The non-OTA producing P. verrucosum inoculated wheat kernels and OTA contaminated wheat kernels subjected to hyperspectral imaging provided different spectral patterns.

Determination of Ochratoxin A in Rye and Rye-Based Products by Fluorescence Polarization Immunoassay

PubMed Central

Lippolis, Vincenzo; Porricelli, Anna C. R.; Cortese, Marina; Zanardi, Sandro; Pascale, Michelangelo

2017-01-01

A rapid fluorescence polarization immunoassay (FPIA) was optimized and validated for the determination of ochratoxin A (OTA) in rye and rye crispbread. Samples were extracted with a mixture of acetonitrile/water (60:40, v/v) and purified by SPE-aminopropyl column clean-up before performing the FPIA. Overall mean recoveries were 86 and 95% for spiked rye and rye crispbread with relative standard deviations lower than 6%. Limits of detection (LOD) of the optimized FPIA was 0.6 μg/kg for rye and rye crispbread, respectively. Good correlations (r > 0.977) were observed between OTA contents in contaminated samples obtained by FPIA and high-performance liquid chromatography (HPLC) with immunoaffinity cleanup used as reference method. Furthermore, single laboratory validation and small-scale collaborative trials were carried out for the determination of OTA in rye according to Regulation 519/2014/EU laying down procedures for the validation of screening methods. The precision profile of the method, cut-off level and rate of false suspect results confirm the satisfactory analytical performances of assay as a screening method. These findings show that the optimized FPIA is suitable for high-throughput screening, and permits reliable quantitative determination of OTA in rye and rye crispbread at levels that fall below the EU regulatory limits. PMID:28954398

Ultrasensitive electrochemical aptasensor for ochratoxin A based on two-level cascaded signal amplification strategy.

PubMed

Yang, Xingwang; Qian, Jing; Jiang, Ling; Yan, Yuting; Wang, Kan; Liu, Qian; Wang, Kun

2014-04-01

Ochratoxin A (OTA) has a number of toxic effects to both humans and animals, so developing sensitive detection method is of great importance. Herein, we describe an ultrasensitive electrochemical aptasensor for OTA based on the two-level cascaded signal amplification strategy with methylene blue (MB) as a redox indicator. In this method, capture DNA, aptamers, and reporter DNA functionalized-gold nanoparticles (GNPs) were immobilized on the electrode accordingly, where GNPs were used as the first-level signal enhancer. To receive the more sensitive response, a larger number of guanine (G)-rich DNA was bound to the GNPs' surface to provide abundant anchoring sites for MB to achieve the second-level signal amplification. By employing this novel strategy, an ~8.5 (±0.3) fold amplification in signal intensity was obtained. Afterward, OTA was added to force partial GNPs/G-rich DNA to release from the sensing interface and thus decreased the electrochemical response. An effective sensing range from 2.5pM to 2.5nM was received with an extremely low detection limit of 0.75 (±0.12) pM. This amplification strategy has the potential to be the main technology for aptamer-based electrochemical biosensor in a variety of fields. Copyright © 2013 Elsevier B.V. All rights reserved.

The effect of chemical treatment on reduction of aflatoxins and ochratoxin A in black and white pepper during washing.

PubMed

Jalili, M; Jinap, S; Son, R

2011-04-01

The effect of 18 different chemicals, which included acidic compounds (sulfuric acid, chloridric acid, phosphoric acid, benzoic acid, citric acid, acetic acid), alkaline compounds (ammonia, sodium bicarbonate, sodium hydroxide, potassium hydroxide, calcium hydroxide), salts (acetate ammonium, sodium bisulfite, sodium hydrosulfite, sodium chloride, sodium sulfate) and oxidising agents (hydrogen peroxide, sodium hypochlorite), on the reduction of aflatoxins B(1), B(2), G(1) and G(2) and ochratoxin A (OTA) was investigated in black and white pepper. OTA and aflatoxins were determined using HPLC after immunoaffinity column clean-up. Almost all of the applied chemicals showed a significant degree of reduction on mycotoxins (p < 0.05). The lowest and highest reduction of aflatoxin B(1), which is the most dangerous aflatoxin, was 20.5% ± 2.7% using benzoic acid and 54.5% ± 2.7% using sodium hydroxide. There was no significant difference between black and white peppers (p < 0.05).

Identification and characterization of the ergochrome gene cluster in the plant pathogenic fungus Claviceps purpurea.

PubMed

Neubauer, Lisa; Dopstadt, Julian; Humpf, Hans-Ulrich; Tudzynski, Paul

2016-01-01

Claviceps purpurea is a phytopathogenic fungus infecting a broad range of grasses including economically important cereal crop plants. The infection cycle ends with the formation of the typical purple-black pigmented sclerotia containing the toxic ergot alkaloids. Besides these ergot alkaloids little is known about the secondary metabolism of the fungus. Red anthraquinone derivatives and yellow xanthone dimers (ergochromes) have been isolated from sclerotia and described as ergot pigments, but the corresponding gene cluster has remained unknown. Fungal pigments gain increasing interest for example as environmentally friendly alternatives to existing dyes. Furthermore, several pigments show biological activities and may have some pharmaceutical value. This study identified the gene cluster responsible for the synthesis of the ergot pigments. Overexpression of the cluster-specific transcription factor led to activation of the gene cluster and to the production of several known ergot pigments. Knock out of the cluster key enzyme, a nonreducing polyketide synthase, clearly showed that this cluster is responsible for the production of red anthraquinones as well as yellow ergochromes. Furthermore, a tentative biosynthetic pathway for the ergot pigments is proposed. By changing the culture conditions, pigment production was activated in axenic culture so that high concentration of phosphate and low concentration of sucrose induced pigment syntheses. This is the first functional analysis of a secondary metabolite gene cluster in the ergot fungus besides that for the classical ergot alkaloids. We demonstrated that this gene cluster is responsible for the typical purple-black color of the ergot sclerotia and showed that the red and yellow ergot pigments are products of the same biosynthetic pathway. Activation of the gene cluster in axenic culture opened up new possibilities for biotechnological applications like the dye production or the development of new pharmaceuticals.

Influence of dietary charcoal on ochratoxin A toxicity in Leghorn chicks.

PubMed Central

Rotter, R G; Frohlich, A A; Marquardt, R R

1989-01-01

The ability of activated charcoal to adsorb ochratoxin A (OA) in vitro and to reduce the toxic effects of OA in vivo when added to the diet of growing Leghorn chicks was studied. Activated charcoal (50 mg) was able to adsorb 90% of the OA (150 micrograms) contained in 10 mL of citrate-phosphate buffer (pH 7.0). When 2 g of a complete chick diet were mixed with OA in buffer, it adsorbed 66% of the OA, while addition of 50 mg of charcoal to this mixture further reduced the concentration of OA to 11.8% of the control, an additional 65% compared to the diet alone. In the first of two feeding studies, charcoal addition of up to 10,000 parts per million (ppm) to diets (6.7% tallow) containing 9.93 mumol (4 ppm) OA kg-1 diet had no effect on OA toxicity. Feed consumption and weight gain, however, were reduced 10 and 20%, respectively, in chicks fed diets which contained 10,000 ppm of charcoal compared to those fed no charcoal. In the second study, reducing dietary tallow to 2% did not alter the effects of OA or charcoal on weight gain and feed to gain ratio, but birds fed OA with 10,000 ppm charcoal had an 8.5% increase in feed consumption. An additional management problem was associated with the propensity of charcoal to blacken the feed, the birds and their environment. Addition of charcoal to OA contaminated diets appeared to be an ineffective method for reducing the toxic effects of OA in growing chicks. PMID:2590872

Ergot alkaloids from endophyte-infected tall fescue decrease reticuloruminal epithelial blood flow and volatile fatty acid absorption from the washed reticulorumen.

PubMed

Foote, A P; Kristensen, N B; Klotz, J L; Kim, D H; Koontz, A F; McLeod, K R; Bush, L P; Schrick, F N; Harmon, D L

2013-11-01

An experiment was conducted to determine if ergot alkaloids affect blood flow to the absorptive surface of the rumen. Steers (n=8) were pair-fed alfalfa cubes and received ground endophyte-infected (Neotyphodium coenophialum) tall fescue (Lolium arundinaceum; E+) seed (0.015 mg ergovaline·kg BW(-1)·d(-1)) or endophyte-free tall fescue (E-) seed via the rumen cannula 2x daily for 7 d at thermoneutral (TN; 22°C) and heat stress (HS; 32°C) conditions. On d 8, the rumen was emptied and rinsed. A buffer containing VFA was incubated in the following sequence: control (CON), 15 μg ergovaline·kg BW(-1) (1×EXT) from a tall fescue seed extract, and 45 μg ergovaline·kg BW(-1) (3×EXT). For each buffer treatment there were two 30-min incubations: a 30-min incubation of a treatment buffer with no sampling followed by an incubation of an identical sampling buffer with the addition of Cr-EDTA and deuterium oxide (D2O). Epithelial blood flow was calculated as ruminal clearance of D2O corrected for influx of physiological water and liquid outflow. Feed intake decreased with dosing E+ seed at HS but not at thermoneutral conditions (TN; P<0.02). Dosing E+ seed decreased serum prolactin (P<0.005) at TN. At HS, prolactin decreased in both groups over the 8-d experiment (P<0.0001), but there was no difference in E+ and E- steers (P=0.33). There was a seed treatment×buffer treatment interaction at TN (P=0.038), indicating that E+ seed treatment decreased reticuloruminal epithelial blood flow at TN during the CON incubation, but the two groups of steers were not different during 1×EXT and 3×EXT (P>0.05). Inclusion of the extract in the buffer caused at least a 50% reduction in epithelial blood flow at TN (P=0.004), but there was no difference between 1×EXT and 3×EXT. There was a seed × buffer treatment interaction at HS (P=0.005), indicating that the reduction of blood flow induced by incubating the extract was larger for steers receiving E- seed than E+ seed. Volatile

Variation in Fumonisin and Ochratoxin Production Associated with Differences in Biosynthetic Gene Content in Aspergillus niger and A. welwitschiae Isolates from Multiple Crop and Geographic Origins

PubMed Central

Susca, Antonia; Proctor, Robert H.; Morelli, Massimiliano; Haidukowski, Miriam; Gallo, Antonia; Logrieco, Antonio F.; Moretti, Antonio

2016-01-01

The fungi Aspergillus niger and A. welwitschiae are morphologically indistinguishable species used for industrial fermentation and for food and beverage production. The fungi also occur widely on food crops. Concerns about their safety have arisen with the discovery that some isolates of both species produce fumonisin (FB) and ochratoxin A (OTA) mycotoxins. Here, we examined FB and OTA production as well as the presence of genes responsible for synthesis of the mycotoxins in a collection of 92 A. niger/A. welwitschiae isolates from multiple crop and geographic origins. The results indicate that (i) isolates of both species differed in ability to produce the mycotoxins; (ii) FB-nonproducing isolates of A. niger had an intact fumonisin biosynthetic gene (fum) cluster; (iii) FB-nonproducing isolates of A. welwitschiae exhibited multiple patterns of fum gene deletion; and (iv) OTA-nonproducing isolates of both species lacked the ochratoxin A biosynthetic gene (ota) cluster. Analysis of genome sequence data revealed a single pattern of ota gene deletion in the two species. Phylogenetic analysis suggest that the simplest explanation for this is that ota cluster deletion occurred in a common ancestor of A. niger and A. welwitschiae, and subsequently both the intact and deleted cluster were retained as alternate alleles during divergence of the ancestor into descendent species. Finally, comparison of results from this and previous studies indicate that a majority of A. niger isolates and a minority of A. welwitschiae isolates can produce FBs, whereas, a minority of isolates of both species produce OTA. The comparison also suggested that the relative abundance of each species and frequency of FB/OTA-producing isolates can vary with crop and/or geographic origin. PMID:27667988

Variation in Fumonisin and Ochratoxin Production Associated with Differences in Biosynthetic Gene Content in Aspergillus niger and A. welwitschiae Isolates from Multiple Crop and Geographic Origins.

PubMed

Susca, Antonia; Proctor, Robert H; Morelli, Massimiliano; Haidukowski, Miriam; Gallo, Antonia; Logrieco, Antonio F; Moretti, Antonio

2016-01-01

The fungi Aspergillus niger and A. welwitschiae are morphologically indistinguishable species used for industrial fermentation and for food and beverage production. The fungi also occur widely on food crops. Concerns about their safety have arisen with the discovery that some isolates of both species produce fumonisin (FB) and ochratoxin A (OTA) mycotoxins. Here, we examined FB and OTA production as well as the presence of genes responsible for synthesis of the mycotoxins in a collection of 92 A. niger/A. welwitschiae isolates from multiple crop and geographic origins. The results indicate that (i) isolates of both species differed in ability to produce the mycotoxins; (ii) FB-nonproducing isolates of A. niger had an intact fumonisin biosynthetic gene (fum) cluster; (iii) FB-nonproducing isolates of A. welwitschiae exhibited multiple patterns of fum gene deletion; and (iv) OTA-nonproducing isolates of both species lacked the ochratoxin A biosynthetic gene (ota) cluster. Analysis of genome sequence data revealed a single pattern of ota gene deletion in the two species. Phylogenetic analysis suggest that the simplest explanation for this is that ota cluster deletion occurred in a common ancestor of A. niger and A. welwitschiae, and subsequently both the intact and deleted cluster were retained as alternate alleles during divergence of the ancestor into descendent species. Finally, comparison of results from this and previous studies indicate that a majority of A. niger isolates and a minority of A. welwitschiae isolates can produce FBs, whereas, a minority of isolates of both species produce OTA. The comparison also suggested that the relative abundance of each species and frequency of FB/OTA-producing isolates can vary with crop and/or geographic origin.

Detection and quantification of ochratoxin A and deoxynivalenol in barley grains by GC-MS and electronic nose.

PubMed

Olsson, J; Börjesson, T; Lundstedt, T; Schnürer, J

2002-02-05

Mycotoxin contamination of cereal grains can be detected and quantified using complex extraction procedures and analytical techniques. Normally, the grain odour, i.e. the presence of non-grain volatile metabolites, is used for quality classification of grain. We have investigated the possibility of using fungal volatile metabolites as indicators of mycotoxins in grain. Ten barley samples with normal odour, and 30 with some kind of off-odour were selected from Swedish granaries. The samples were evaluated with regard to moisture content, fungal contamination, ergosterol content, and levels of ochratoxin A (OA) and deoxynivalenol (DON). Volatile compounds were also analysed using both an electronic nose and gas chromatography combined with mass spectrometry (GC-MS). Samples with normal odour had no detectable ochratoxin A and average DON contents of 16 microg kg(-1) (range 0-80), while samples with off-odour had average OA contents of 76 microg kg(-1) (range 0-934) and DON contents of 69 microg kg(-1) (range 0-857). Data were evaluated by multivariate data analysis using projection methods such as principal component analysis (PCA) and partial least squares (PLS). The results show that it was possible to classify the OA level as below or above the maximum limit of 5 microg kg(-1) cereal grain established by the Swedish National Food Administration, and that the DON level could be estimated using PLS. Samples with OA levels below 5 microg kg(-1) had higher concentration of aldehydes (nonanal, 2-hexenal) and alcohols (1-penten-3-ol, 1-octanol). Samples with OA levels above 5 microg kg(-1) had higher concentrations of ketones (2-hexanone, 3-octanone). The GC-MS system predicted OA concentrations with a higher accuracy than the electronic nose, since the GC-MS misclassified only 3 of 37 samples and the electronic nose 7 of 37 samples. No correlation was found between odour and OA level, as samples with pronounced or strong off-odours had OA levels both below and above 5

MALDI-Imaging Mass Spectrometry of Ochratoxin A and Fumonisins in Mold-Infected Food.

PubMed

Hickert, Sebastian; Cramer, Benedikt; Letzel, Matthias C; Humpf, Hans-Ulrich

2016-09-06

Mycotoxins are toxic secondary metabolites produced by various fungi. Their distribution within contaminated material is of high interest to obtain insight into infection mechanisms and the possibility of reducing contamination during food processing. Various vegetable foodstuffs were infected with fungi of the genera Fusarium and Aspergillus. The localization of the produced mycotoxins was studied by matrix assisted laser desorption ionization time of flight imaging mass spectrometry (MALDI-MSI) of cryosections obtained from infected material. The results were confirmed by HPLC-electrospray ionization triple quadrupole mass spectrometry (HPLC/MS/MS). The mycotoxins ochratoxin A (OTA) and fumonisins of the B- and C-series (FB 1 , FB 2 , FB 3 , FB 4 , FC 2/3 , and FC 4 ) as well as partially hydrolyzed fumonisins (pHFB 1 , pHFB 2 , pHFB 3 , pHFC 1 , and pHFC 2/3 ) could successfully be detected by MALDI-IMS in mold-infested foodstuffs. The toxins are distributed differently in the material: OTA is co-localized with visible fungal spoilage while fumonisins could be detected throughout the whole sample. This work shows the applicability of MALDI-Imaging Mass Spectrometry (MALDI-MSI) to mycotoxin analysis. It has been demonstrated that the analyzed mycotoxins are differently distributed within moldy foodstuffs. These findings show the potential of MALDI-MSI for the localization of these hazardous compounds in various plant tissues. This article is protected by copyright. All rights reserved.

Evaluation of extraction methods for ochratoxin A detection in cocoa beans employing HPLC.

PubMed

Mishra, Rupesh K; Catanante, Gaëlle; Hayat, Akhtar; Marty, Jean-Louis

2016-01-01

Cocoa is an important ingredient for the chocolate industry and for many food products. However, it is prone to contamination by ochratoxin A (OTA), which is highly toxic and potentially carcinogenic to humans. In this work, four different extraction methods were tested and compared based on their recoveries. The best protocol was established which involves an organic solvent-free extraction method for the detection of OTA in cocoa beans using 1% sodium hydrogen carbonate (NaHCO3) in water within 30 min. The extraction method is rapid (as compared with existing methods), simple, reliable and practical to perform without complex experimental set-ups. The cocoa samples were freshly extracted and cleaned-up using immunoaffinity column (IAC) for HPLC analysis using a fluorescence detector. Under the optimised condition, the limit of detection (LOD) and limit of quantification (LOQ) for OTA were 0.62 and 1.25 ng ml(-1) respectively in standard solutions. The method could successfully quantify OTA in naturally contaminated samples. Moreover, good recoveries of OTA were obtained up to 86.5% in artificially spiked cocoa samples, with a maximum relative standard deviation (RSD) of 2.7%. The proposed extraction method could determine OTA at the level 1.5 µg kg(-)(1), which surpassed the standards set by the European Union for cocoa (2 µg kg(-1)). In addition, an efficiency comparison of IAC and molecular imprinted polymer (MIP) column was also performed and evaluated.

Amplified impedimetric immunosensor based on instant catalyst for sensitive determination of ochratoxin A.

PubMed

Tang, Juan; Huang, Yapei; Zhang, Cengceng; Liu, Huiqiong; Tang, Dianping

2016-12-15

A new impedimetric immunosensor for the fast determination of ochratoxin A (OTA) in food samples was developed based on the instant catalyst as enhancer. Initially, the signal tags were prepared via co-immobilization of anti-OTA antibody and amine-terminated dendrimer (PAMAM) on the graphene oxide nanosheets through the covalent interaction, which were utilized as a good platform for combining manganese ion (anti-OTA-GO-PAMAM-Mn(2+)). Upon target OTA introduction, a competitive-type immunoreaction was implemented between the analyte and the immobilized OTA-BSA on the electrode for the anti-OTA antibody on the graphene oxide nanosheets labels. After a competitive immunoassay format, the anti-OTA-GO-PAMAM-Mn(2+) were captured onto the electrode surface, which could induce the in situ formation of MnO2via classical redox reaction between Mn(2+) and KMnO4 on the immunesensing platform. Moreover, the generated MnO2 nanoparticles act as efficient catalyst could catalyze the 4-chloro-1-naphthol (4-CN) oxidation without H2O2 to generate an insoluble precipitation on the platform. Under the optimal conditions, the instant catalyst based impedimetric immunosensor displayed a wide dynamic working range between 0.1pgmL(-1) and 30ngmL(-1). The detection limit (LOD) of the assay was 0.055pgmL(-1). The developed method exhibited high selectivity and can be used for the determination of OTA in real red wine samples. Copyright © 2016 Elsevier B.V. All rights reserved.

Health risk assessment of ochratoxin A for all age-sex strata in a market economy.

PubMed

Kuiper-Goodman, T; Hilts, C; Billiard, S M; Kiparissis, Y; Richard, I D K; Hayward, S

2010-02-01

In order to manage risk of ochratoxin A (OTA) in foods, we re-evaluated the tolerable daily intake (TDI), derived the negligible cancer risk intake (NCRI), and conducted a probabilistic risk assessment. A new approach was developed to derive 'usual' probabilistic exposure in the presence of highly variable occurrence data, such as encountered with low levels of OTA. Canadian occurrence data were used for various raw food commodities or finished foods and were combined with US Department of Agriculture (USDA) food consumption data, which included data on infants and young children. Both variability and uncertainty in input data were considered in the resulting exposure estimates for various age/sex strata. Most people were exposed to OTA on a daily basis. Mean adjusted exposures for all age-sex groups were generally below the NCRI of 4 ng OTA kg bw(-1), except for 1-4-year-olds as a result of their lower body weight. For children, the major contributors of OTA were wheat-based foods followed by oats, rice, and raisins. Beer, coffee, and wine also contributed to total OTA exposure in older individuals. Predicted exposure to OTA decreased when European Commission maximum limits were applied to the occurrence data. The impact on risk for regular eaters of specific commodities was also examined.

Anorexia Induction by the Trichothecene Deoxynivalenol (Vomitoxin) Is Mediated by the Release of the Gut Satiety Hormone Peptide YY

PubMed Central

Pestka, James J.

2012-01-01

Consumption of deoxynivalenol (DON), a trichothecene mycotoxin known to commonly contaminate grain-based foods, suppresses growth of experimental animals, thus raising concerns over its potential to adversely affect young children. Although this growth impairment is believed to result from anorexia, the initiating mechanisms for appetite suppression remain unknown. Here, we tested the hypothesis that DON induces the release of satiety hormones and that this response corresponds to the toxin’s anorectic action. Acute ip exposure to DON had no effect on plasma glucagon-like peptide-1, leptin, amylin, pancreatic polypeptide, gastric inhibitory peptide, or ghrelin; however, the toxin was found to robustly elevate peptide YY (PYY) and cholecystokinin (CCK). Specifically, ip exposure to DON at 1 and 5mg/kg bw induced PYY by up to 2.5-fold and CCK by up to 4.1-fold. These responses peaked within 15–120min and lasted up to 120min (CCK) and 240min (PPY), corresponding with depressed rates of food intake. Direct administration of exogenous PYY or CCK similarly caused reduced food intake. Food intake experiments using the NPY2 receptor antagonist BIIE0246 and the CCK1A receptor antagonist devazepide, individually, suggested that PYY mediated DON-induced anorexia but CCK did not. Orolingual exposure to DON induced plasma PYY and CCK elevation and anorexia comparable with that observed for ip exposure. Taken together, these findings suggest that PYY might be one critical mediator of DON-induced anorexia and, ultimately, growth suppression. PMID:22903826

Anorexia induction by the trichothecene deoxynivalenol (vomitoxin) is mediated by the release of the gut satiety hormone peptide YY.

PubMed

Flannery, Brenna M; Clark, Erica S; Pestka, James J

2012-12-01

Consumption of deoxynivalenol (DON), a trichothecene mycotoxin known to commonly contaminate grain-based foods, suppresses growth of experimental animals, thus raising concerns over its potential to adversely affect young children. Although this growth impairment is believed to result from anorexia, the initiating mechanisms for appetite suppression remain unknown. Here, we tested the hypothesis that DON induces the release of satiety hormones and that this response corresponds to the toxin's anorectic action. Acute ip exposure to DON had no effect on plasma glucagon-like peptide-1, leptin, amylin, pancreatic polypeptide, gastric inhibitory peptide, or ghrelin; however, the toxin was found to robustly elevate peptide YY (PYY) and cholecystokinin (CCK). Specifically, ip exposure to DON at 1 and 5mg/kg bw induced PYY by up to 2.5-fold and CCK by up to 4.1-fold. These responses peaked within 15-120 min and lasted up to 120 min (CCK) and 240 min (PPY), corresponding with depressed rates of food intake. Direct administration of exogenous PYY or CCK similarly caused reduced food intake. Food intake experiments using the NPY2 receptor antagonist BIIE0246 and the CCK1A receptor antagonist devazepide, individually, suggested that PYY mediated DON-induced anorexia but CCK did not. Orolingual exposure to DON induced plasma PYY and CCK elevation and anorexia comparable with that observed for ip exposure. Taken together, these findings suggest that PYY might be one critical mediator of DON-induced anorexia and, ultimately, growth suppression.

Use of enzyme-linked immunosorbent assay to screen for aflatoxins, ochratoxin A, and deoxynivalenol in dry pet foods.

PubMed

Okuma, Tara A; Huynh, Thu P; Hellberg, Rosalee S

2018-03-01

The objective of this study was to perform a market survey on dry pet foods using enzyme-linked immunosorbent assay (ELISA) to detect total aflatoxins (AFs), ochratoxin A (OTA), and deoxynivalenol (DON). Pet food products (n = 58) marketed for dogs, cats, birds, and rabbits were tested in duplicate with ELISA, and results above the limit of quantitation were confirmed using liquid chromatography tandem mass spectrometry (LC-MS/MS). OTA was detected in one product (rabbit food) and AFs were detected in two products (one dog treat and one bird treat). In contrast, DON was detected in the majority (74%) of products tested. Bird and rabbit products were the most affected by DON, with levels above 0.5 μg/g in 50 and 80% of samples, respectively. One rabbit sample tested positive for both OTA and DON. Overall, the findings of this study revealed a low incidence of AFs and OTA in commercial pet food. Although DON was detected in numerous products, the levels were well below those associated with acute toxic effects.

Reduction of Ochratoxin A in Oat Flakes by Twin-Screw Extrusion Processing.

PubMed

Lee, Hyun Jung; Dahal, Samjhana; Perez, Enrique Garcia; Kowalski, Ryan Joseph; Ganjyal, Girish M; Ryu, Dojin

2017-10-01

Ochratoxin A (OTA) is one of the most important mycotoxins owing to its widespread occurrence and toxicity, including nephrotoxicity and potential carcinogenicity to humans. OTA has been detected in a wide range of agricultural commodities, including cereal grains and their processed products. In particular, oat-based products show a higher incidence and level of contamination. Extrusion cooking is widely used in the manufacturing of breakfast cereals and snacks and may reduce mycotoxins to varying degrees. Hence, the effects of extrusion cooking on the stability of OTA in spiked (100 μg/kg) oat flake was investigated by using a laboratory-scale twin-screw extruder with a central composite design. Factors examined were moisture content (20, 25, and 30% dry weight basis), temperature (140, 160, and 180°C), screw speed (150, 200, and 250 rpm), and die size (1.5, 2, and 3 mm). Both nonextruded and extruded samples were analyzed for reductions of OTA by high-performance liquid chromatography, coupled with fluorescence detection. The percentage of reductions in OTA in the contaminated oat flakes upon extrusion processing were in the range of 0 to 28%. OTA was partially stable during extrusion, with only screw speed and die size having significant effect on reduction (P < 0.005). The highest reduction of 28% was achieved at 180°C, 20% moisture, 250 rpm screw speed, and a 3-mm die with 193 kJ/kg specific mechanical energy. According to the central composite design analyses, up to 28% of OTA can be reduced by a combination of 162°C, 30% moisture, and 221 rpm, with a 3-mm die.

Proteome analysis of the fungus Aspergillus carbonarius under ochratoxin A producing conditions.

PubMed

Crespo-Sempere, A; Gil, J V; Martínez-Culebras, P V

2011-06-30

Aspergillus carbonarius is an important ochratoxin A producing fungus that is responsible for mycotoxin contamination of grapes and wine. In this study, the proteomes of highly (W04-40) and weakly (W04-46) OTA-producing A. carbonarius strains were compared to identify proteins that may be involved in OTA biosynthesis. Protein samples were extracted from two biological replicates and subjected to two dimensional gel electrophoresis analysis and mass spectrometry. Expression profile comparison (PDQuest software), revealed 21 differential spots that were statistically significant and showed a two-fold change in expression, or greater. Among these, nine protein spots were identified by MALDI-MS/MS and MASCOT database and twelve remain unidentified. Of the identified proteins, seven showed a higher expression in strain W04-40 (high OTA producer) and two in strain W04-46 (low OTA producer). Some of the identified amino acid sequences shared homology with proteins involved in regulation, amino acid metabolism, oxidative stress and sporulation. It is worth noting the presence of a protein with 126.5 fold higher abundance in strain W04-40 showing homology with protein CipC, a protein with unknown function related with pathogenesis and mycotoxin production by some authors. Variations in protein expression were also further investigated at the mRNA level by real-time PCR analysis. The mRNA expression levels from three identified proteins including CipC showed correlation with protein expression levels. This study represents the first proteomic analysis for a comparison of two A. carbonarius strains with different OTA production and will contribute to a better understanding of the molecular events involved in OTA biosynthesis. Copyright © 2011 Elsevier B.V. All rights reserved.

Ochratoxin A in raisins and currants: basic extraction procedure used in two small marketing surveys of the occurrence and control of the heterogeneity of the toxins in samples.

PubMed

Möller, T E; Nyberg, M

2003-11-01

A basic extraction procedure for analysis of ochratoxin A (OTA) in currants and raisins is described, as well as the occurrence of OTA and a control of heterogeneity of the toxin in samples bought for two small marketing surveys 1999/2000 and 2001/02. Most samples in the surveys were divided into two subsamples that were individually prepared as slurries and analysed separately. The limit of quantification for the method was estimated as 0.1 microg kg(-1) and recoveries of 85, 90 and 115% were achieved in recovery experiments at 10, 5 and 0.1 microg kg(-1), respectively. Of all 118 subsamples analysed in the surveys, 96 (84%) contained ochratoxin A at levels above the quantification level and five samples (4%) contained more than the European Community legislation of 10 microg kg(-1). The OTA concentrations found in the first survey were in the range < 0.1-19.0 microg kg(-1) with a median concentration of 0.9 microg kg(-1). In the 2001/02 study, the range was < 0.1-34.6 microg kg(-1) with a median of 0.2 microg kg(-1). Big differences were often achieved between individual subsamples of the original sample, which indicate a wide heterogeneous distribution of the toxin. Data from the repeatability test as well as recovery experiments from the same slurries showed that preparation of slurries as described here seemed to give a homogeneous and representative sample. The extraction with the basic sodium bicarbonate-methanol mixture used in the surveys gave similar or somewhat higher OTA values on some samples tested in a comparison with a weak phosphoric acid water-methanol extraction mixture.

Experimentally determined soil organic matter-water sorption coefficients for different classes of natural toxins and comparison with estimated numbers.

PubMed

Schenzel, Judith; Goss, Kai-Uwe; Schwarzenbach, René P; Bucheli, Thomas D; Droge, Steven T J

2012-06-05

Although natural toxins, such as mycotoxins or phytoestrogens are widely studied and were recently identified as micropollutants in the environment, many of their environmentally relevant physicochemical properties have not yet been determined. Here, the sorption affinity to Pahokee peat, a model sorbent for soil organic matter, was investigated for 29 mycotoxins and two phytoestrogens. Sorption coefficients (K(oc)) were determined with a dynamic HPLC-based column method using a fully aqueous mobile phase with 5 mM CaCl(2) at pH 4.5. Sorption coefficients varied from less than 10(0.7) L/kg(oc) (e.g., all type B trichothecenes) to 10(4.0) L/kg(oc) (positively charged ergot alkaloids). For the neutral compounds the experimental sorption data set was compared with predicted sorption coefficients using various models, based on molecular fragment approaches (EPISuite's KOCWIN or SPARC), poly parameter linear free energy relationship (pp-LFER) in combination with predicted descriptors, and quantum-chemical based software (COSMOtherm)). None of the available models was able to adequately predict absolute K(oc) numbers and relative differences in sorption affinity for the whole set of neutral toxins, largely because mycotoxins exhibit highly complex structures. Hence, at present, for such compounds fast and consistent experimental techniques for determining sorption coefficients, as the one used in this study, are required.

Effect of gaseous ozone treatments on DON, microbial contaminants and technological parameters of wheat and semolina.

PubMed

Piemontese, Luca; Messia, Maria Cristina; Marconi, Emanuele; Falasca, Luisa; Zivoli, Rosanna; Gambacorta, Lucia; Perrone, Giancarlo; Solfrizzo, Michele

2018-04-01

Deoxynivalenol (DON) is an important mycotoxin produced by several species of Fusarium. It occurs often in wheat grain and is frequently associated with significant levels of its modified form DON-3-glucoside (DON-3-Glc). Ozone (O 3 ) is a powerful disinfectant and oxidant, classified as GRAS (Generally Recognised As Safe), that reacts easily with specific compounds including the mycotoxins aflatoxins, ochratoxin A, trichothecenes and zearalenone. It degrades DON in aqueous solution and can be effective for decontamination of grain. This study reports the efficacy of gaseous ozone treatments in reducing DON, DON-3-Glc, bacteria, fungi and yeasts in naturally contaminated durum wheat. A prototype was used to dispense ozone continuously and homogeneously at different concentrations and exposure time, in 2 kg aliquots of durum wheat. The optimal conditions, which do not affect chemical and rheological parameters of durum wheat, semolina and pasta, were identified (55 g O 3  h -1 for 6 h). The measured mean reductions of DON and DON-3-Glc in ozonated wheat were 29% and 44%, respectively. Ozonation also produced a significant (p < 0.05) reduction of total count (CFU/g) of bacteria, fungi and yeasts in wheat grains.

Occurrence, Toxicity, and Analysis of Major Mycotoxins in Food

PubMed Central

Alshannaq, Ahmad; Yu, Jae-Hyuk

2017-01-01

Mycotoxins are toxic secondary metabolites produced by certain filamentous fungi (molds). These low molecular weight compounds (usually less than 1000 Daltons) are naturally occurring and practically unavoidable. They can enter our food chain either directly from plant-based food components contaminated with mycotoxins or by indirect contamination from the growth of toxigenic fungi on food. Mycotoxins can accumulate in maturing corn, cereals, soybeans, sorghum, peanuts, and other food and feed crops in the field and in grain during transportation. Consumption of mycotoxin-contaminated food or feed can cause acute or chronic toxicity in human and animals. In addition to concerns over adverse effects from direct consumption of mycotoxin-contaminated foods and feeds, there is also public health concern over the potential ingestion of animal-derived food products, such as meat, milk, or eggs, containing residues or metabolites of mycotoxins. Members of three fungal genera, Aspergillus, Fusarium, and Penicillium, are the major mycotoxin producers. While over 300 mycotoxins have been identified, six (aflatoxins, trichothecenes, zearalenone, fumonisins, ochratoxins, and patulin) are regularly found in food, posing unpredictable and ongoing food safety problems worldwide. This review summarizes the toxicity of the six mycotoxins, foods commonly contaminated by one or more of them, and the current methods for detection and analysis of these mycotoxins. PMID:28608841

Review on Mycotoxin Issues in Ruminants: Occurrence in Forages, Effects of Mycotoxin Ingestion on Health Status and Animal Performance and Practical Strategies to Counteract Their Negative Effects

PubMed Central

Gallo, Antonio; Giuberti, Gianluca; Frisvad, Jens C.; Bertuzzi, Terenzio; Nielsen, Kristian F.

2015-01-01

Ruminant diets include cereals, protein feeds, their by-products as well as hay and grass, grass/legume, whole-crop maize, small grain or sorghum silages. Furthermore, ruminants are annually or seasonally fed with grazed forage in many parts of the World. All these forages could be contaminated by several exometabolites of mycotoxigenic fungi that increase and diversify the risk of mycotoxin exposure in ruminants compared to swine and poultry that have less varied diets. Evidence suggests the greatest exposure for ruminants to some regulated mycotoxins (aflatoxins, trichothecenes, ochratoxin A, fumonisins and zearalenone) and to many other secondary metabolites produced by different species of Alternaria spp. (e.g., AAL toxins, alternariols, tenuazonic acid or 4Z-infectopyrone), Aspergillus flavus (e.g., kojic acid, cyclopiazonic acid or β-nitropropionic acid), Aspergillus fuminatus (e.g., gliotoxin, agroclavine, festuclavines or fumagillin), Penicillium roqueforti and P. paneum (e.g., mycophenolic acid, roquefortines, PR toxin or marcfortines) or Monascus ruber (citrinin and monacolins) could be mainly related to forage contamination. This review includes the knowledge of mycotoxin occurrence reported in the last 15 years, with special emphasis on mycotoxins detected in forages, and animal toxicological issues due to their ingestion. Strategies for preventing the problem of mycotoxin feed contamination under farm conditions are discussed. PMID:26274974

21 CFR 310.531 - Drug products containing active ingredients offered over-the-counter (OTC) for the treatment of...

Code of Federal Regulations, 2013 CFR

2013-04-01

... subnitrate, calomel, camphor, cholesterol, ergot fluid extract, hexachlorophene, ichthammol, isobutamben... aminacrine hydrochloride, bismuth subnitrate, calomel, camphor, cholesterol, ergot fluid extract...

21 CFR 310.531 - Drug products containing active ingredients offered over-the-counter (OTC) for the treatment of...

Code of Federal Regulations, 2011 CFR

2011-04-01

... subnitrate, calomel, camphor, cholesterol, ergot fluid extract, hexachlorophene, ichthammol, isobutamben... aminacrine hydrochloride, bismuth subnitrate, calomel, camphor, cholesterol, ergot fluid extract...

21 CFR 310.531 - Drug products containing active ingredients offered over-the-counter (OTC) for the treatment of...

Code of Federal Regulations, 2010 CFR

2010-04-01

... subnitrate, calomel, camphor, cholesterol, ergot fluid extract, hexachlorophene, ichthammol, isobutamben... aminacrine hydrochloride, bismuth subnitrate, calomel, camphor, cholesterol, ergot fluid extract...