zearalenone and beauvericin

Cell cycle progression and programmed cell death are imposed by pathological stimuli of extrinsic or intrinsic including the exposure to neurotoxins, oxidative stress and DNA damage. All can cause abrupt or delayed cell death, inactivate normal cell survival or cell death networks. Nevertheless, the mechanisms of the neuronal cell death are unresolved. One of the cell deaths triggers which have been wildly studied, correspond to mycotoxins produced by Fusarium species, which have been demonstrated cytotoxicity and neurotoxicity through impairing cell proliferation, gene expression and induction of oxidative stress. The aim of present study was to analyze the cell cycle progression and cell death pathway by flow cytometry in undifferentiated SH-SY5Y neuronal cells exposed to α-zearalenol (α-ZEL), β-zearalenol (β-ZEL) and beauvericin (BEA) over 24 h and 48 h individually and combined at the following concentration ranges: from 1.56 to 12.5 μM for α-ZEL and β-ZEL, from 0.39 to 2.5 μM for BEA, from 1.87 to 25 μM for binary combinations and from 3.43 to 27.5 μM for tertiary combination. Alterations in cell cycle were observed remarkably for β-ZEL at the highest concentration in all treatments where engaged (β-ZEL, β-ZEL + BEA and β-ZEL + α-ZEL), for both 24 h and 48 h. by activating the cell proliferation in G0/G1 phase (up to 43.6 %) and causing delays or arrests in S and G2/M phases (up to 19.6 %). Tertiary mixtures revealed increases of cell proliferation in subG0 phase by 4-folds versus control. Similarly, for cell death among individual treatments β-ZEL showed a significant growth in early apoptotic cells population at the highest concentration assayed as well as for all combination treatments where β-ZEL was involved, in both early apoptotic and apoptotic/necrotic cell death pathways.

Topics: Apoptosis; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Depsipeptides; Estrogens, Non-Steroidal; Humans; Mycotoxins; Zearalenone

The co-presence of mycotoxins from fungi of the genus Fusarium is a common fact in raw food and food products, as trace levels of them or their metabolites can be detected, unless safety practices during manufacturing are carried out. Zearalenone (ZEA), its metabolites α-zearalenol (α-ZEL) and β-zearalenol (β-ZEL) and, beauvericin (BEA) are co/present in cereals, fruits or their products which is a mixture that consumer are exposed and never evaluated in neuronal cells. In this study the role of oxidative stress and intracellular defense systems was assessed by evaluating reactive oxygen species (ROS) generation and glutathione (GSH) ratio activity in a human neuroblastoma cell line, SH-SY5Y cells, treated individually and combined with α-ZEL, β-ZEL and BEA. It was further examined the expression of genes involved in cell apoptosis (CASP3, BAX, BCL2) and receptors of (endogenous or exogenous) estrogens (ERβ and GPER1), by RT-PCR in those same conditions. These results demonstrated elevated ROS levels in combinations where α-ZEL was involved (2.8- to 8-fold compared to control); however, no significant difference in ROS levels were detected when single mycotoxin was tested. Also, the results revealed a significant increase in GSH/GSSG ratio at all concentrations after 24 h. Expression levels of CASP3 and BAX were up regulated by α-ZEL while CASP3 and BCL2 were down regulated by β-ZEL, revealing how ZEA´s metabolites can induce the expression of cell apoptosis genes. However, BEA down-regulated the expression of BCL2. Moreover, β-ZEL + BEA was the only combination treatment which was able to down regulate the levels of cell apoptosis gene expression. Relying to our findings, α-ZEL, β-ZEL and BEA, induce injury in SH-SY5Y cells elevating oxidative stress levels, disturbing the antioxidant activity role of glutathione system and finally, causing disorder in the expressions and activities of the related apoptotic cell death genes.

Topics: Apoptosis; Caspase 3; Cell Culture Techniques; Cell Line, Tumor; Depsipeptides; Gene Expression; Glutathione; Humans; Oxidation-Reduction; Oxidative Stress; Reactive Oxygen Species; Up-Regulation; Zearalenone; Zeranol

Over a 4-year period (2010-13), a survey aiming at determining the occurrence of Fusarium spp. and their relations to mycotoxins in mature grains took place in southern Belgium. The most prevalent species were F. graminearum, F. avenaceum, F. poae and F. culmorum, with large variations between years and locations. An even proportion of mating type found for F. avenaceum, F. culmorum, F. cerealis and F. tricinctum is usually a sign of ongoing sexual recombination. In contrast, an unbalanced proportion of mating type was found for F. poae and no MAT1-2 allele was present in the F. langsethiae population. Genetic chemotyping indicates a majority of deoxynivalenol (DON)-producing strains in F. culmorum (78%, all 3-ADON producers) and F. graminearum (95%, mostly 15-ADON producers), while all F. cerealis strains belong to the nivalenol (NIV) chemotype. Between 2011 and 2013, DON, NIV, enniatins (ENNs) and moniliformin (MON) were found in each field in various concentrations. By comparison, beauvericin (BEA) was scarcely detected and T-2 toxin, zearalenone and α- and β-zearalenols were never detected. Principal component analysis revealed correlations of DON with F. graminearum, ENNs and MON with F. avenaceum and NIV with F. culmorum, F. cerealis and F. poae. BEA was associated with the presence of F. tricinctum and, to a lesser extent, with the presence of F. poae. The use of genetic chemotype data revealed that DON concentrations were mostly influenced by DON-producing strains of F. graminearum and F. culmorum, whereas the concentrations of NIV were influenced by the number of NIV-producing strains of both species added to the number of F. cerealis and F. poae strains. This study emphasises the need to pay attention to less-studied Fusarium spp. for future Fusarium head blight management strategies, as they commonly co-occur in the field and are associated with a broad spectrum of mycotoxins.

Topics: Belgium; Depsipeptides; DNA, Fungal; Edible Grain; Food Contamination; Fusarium; Genes, Mating Type, Fungal; Humans; Mycotoxins; Principal Component Analysis; Trichothecenes; Zearalenone

Wheat is often infected by Fusarium species producing mycotoxins, which may pose health risks to humans and animals. Deoxynivalenol (DON) is the most important Fusarium toxin in Swedish wheat and has previously been shown to be produced mainly by Fusarium graminearum. However, less is known about the co-occurrence of DON and F. graminearum with other toxins and Fusarium species in Sweden. This study examined the distribution of the most important toxigenic Fusarium species and their toxins in winter wheat (2009 and 2011) and spring wheat (2010 and 2011). DNA from seven species was quantified with qPCR and the toxin levels were quantified with a multitoxin analysis method based on liquid chromatography/electrospray ionisation-tandem mass spectrometry (HPLC/ESI-MS/MS). The method enabled detection of many fungal metabolites, including DON, zearalenone (ZEA), nivalenol (NIV), T-2 toxin, HT-2 toxins, moniliformin (MON), beauvericin (BEA), and enniatins (ENNs). It was found that Fusarium poae and Fusarium avenaceum were present in almost all samples. Other common Fusarium species were F. graminearum and F. culmorum, present in more than 70% of samples. Several species occurred at lower DNA levels in 2011 than in other years, but the reverse was true for F. graminearum and Fusarium langsethiae. The most prevalent toxins were ENNs, present in 100% of samples. DON was also common, especially in spring wheat, whereas ZEA and NIV were common in 2009 and in winter wheat, but less common in 2011 and in spring wheat. Only three samples of spring wheat contained T-2 or HT-2 above LOQ. Annual mean levels of several mycotoxins were significantly lower in 2011 than in other years, but the reverse applied for DON. The strongest correlations between mycotoxin and Fusarium DNA levels were found between F. avenaceum and ENNs (r(2) = 0.67) and MON (r(2) = 0.62), and F. graminearum and DON (r(2) = 0.74). These results show that several Fusarium species and toxins co-occur in wheat. The highest toxin levels were detected in spring wheat and DON and ENNs, the latter belonging to the group of so called "emerging toxins", which were the most prevalent toxins and those occurring at the highest levels.

Topics: Chromatography, High Pressure Liquid; Cyclobutanes; Depsipeptides; DNA, Fungal; Food Contamination; Fusarium; Real-Time Polymerase Chain Reaction; Sweden; T-2 Toxin; Tandem Mass Spectrometry; Trichothecenes; Triticum; Zearalenone

Fusarium moulds frequently contaminate oats and other cereals world-wide, including those grown in Northern Europe. To investigate the presence of toxigenic Fusarium species and their toxins in oats, samples were taken during 2010 and 2011 in three geographical regions of Sweden (east, west, south). The samples were analysed by real-time PCR for the specific infection level of seven Fusarium species associated with oats and other cereals (Fusarium poae, Fusarium graminearum, Fusarium langsethiae, Fusarium culmorum, Fusarium tricinctum, Fusarium sporotrichioides and Fusarium avenaceum) and with a multi-mycotoxin method based on liquid chromatography/electrospray ionisation-tandem mass spectrometry (HPLC/ESI-MS/MS) for the detection of many fungal metabolites, including deoxynivalenol (DON), zearalenone (ZEA), nivalenol (NIV), T-2 toxin, HT-2 toxins, moniliformin (MON), beauvericin (BEA) and enniatins (ENNs). Most samples contained at least four of the seven Fusarium species analysed and F. poae, F. langsethiae and F. avenaceum were present in approximately 90-100% of all samples. The most common toxins detected were DON, NIV, BEA and ENNs, which were present in more than 90% of samples. Most Fusarium species and their toxins occurred in higher concentrations in 2010 than in 2011, with the exception of DON and its main producer F. graminearum. Significant regional differences were detected for some moulds and mycotoxins, with higher levels of F. graminearum, DON and ZEA in western Sweden than in the east (P<0.05) and higher levels of F. tricinctum and MON in the south (P<0.05). Correlation analysis showed significant correlations between many Fusarium species and toxin levels. For example, F. tricinctum was significantly correlated to F. avenaceum (r = 0.72, P<0.001), DON to ZEA (r = 0.52, P<0.001), DON to F. graminearum (r = 0.77, P<0.001) and the sum of T-2 and HT-2 to F. langsethiae (r = 0.77, P<0.001). The multi-toxin approach employed allowed simultaneous detection of many Fusarium mycotoxins in each sample. In combination with real-time PCR analysis of seven toxigenic Fusarium spp., the results gave an overall picture of the presence of Fusarium and their toxins in Swedish oats and revealed significant annual and regional differences. This is the first study of the so-called emerging mycotoxins (e.g., ENNs, MON and BEA) in oats grown in Sweden.

Topics: Avena; Chromatography, High Pressure Liquid; Cyclobutanes; Depsipeptides; DNA, Fungal; Edible Grain; Food Contamination; Fusarium; Geography; Real-Time Polymerase Chain Reaction; Sweden; T-2 Toxin; Tandem Mass Spectrometry; Trichothecenes; Zearalenone

Mycotoxins are known to affect the health of humans and husbandry animals. In contrast to wheat grains used for food and feed, whole wheat plants are rarely analysed for mycotoxins, although contaminated straw could additionally expose animals to these toxic compounds. Since the entire wheat plant may also act as source of mycotoxins emitted into the environment, an analytical method was developed, optimised and validated for the analysis of 28 different mycotoxins in above-ground material from whole wheat plants. The method comprises solid-liquid extraction and a clean-up step using a Varian Bond Elut Mycotoxin(®) cartridge, followed by liquid chromatography with electrospray ionisation and triple quadrupole mass spectrometry. Total method recoveries for 26 out of 28 compounds were between 69 and 122% and showed limits of detection from 1 to 26 ng/gdry weight (dw). The overall repeatability for all validated compounds was on average 7%, and their mean ion suppression 65%. Those rather high matrix effects made it necessary to use matrix-matched calibrations to quantify mycotoxins within whole wheat plants. The applicability of this method is illustrated with data from a winter wheat test field to examine the risks of environmental contamination by toxins following artificial inoculation separately with four different Fusarium species. The selected data originate from samples of a part of the field which was inoculated with Fusarium crookwellense. In the wheat samples, various trichothecenes (3-acetyl-deoxynivalenol, deoxynivalenol, diacetoxyscirpenol, fusarenone-X, nivalenol, HT-2 toxin, and T-2 toxin) as well as beauvericin and zearalenone were identified with concentrations ranging from 32 ng/gdw to 12 × 10(3) ng/gdw.

Topics: Chromatography, Liquid; Depsipeptides; Edible Grain; Fusarium; Mycotoxins; Plant Diseases; Spectrometry, Mass, Electrospray Ionization; Trichothecenes; Triticum; Zearalenone

Most recent information on the occurrence of Fusarium Head Blight species and related mycotoxins in wheat grown in the Netherlands dates from 2001. This aim of this study was to investigate the incidence and levels of Fusarium Head Blight species and Fusarium mycotoxins, as well as their possible relationships, in winter wheat cultivated in the Netherlands in 2009. Samples were collected from individual fields of 88 commercial wheat growers. Samples were collected at harvest from 86 fields, and 2 weeks before the expected harvest date from 21 fields. In all, 128 samples, the levels of each of seven Fusarium Head Blight species and of 12 related mycotoxins were quantified. The results showed that F. graminearum was the most frequently observed species at harvest, followed by F. avenaceum and M. nivale. In the pre-harvest samples, only F. graminearum and M. nivale were relevant. The highest incidence and concentrations of mycotoxins were found for deoxynivalenol, followed by zearalenone and beauvericin, both pre-harvest and at harvest. Other toxins frequently found--for the first time in the Netherlands--included T-2 toxin, HT-2 toxin, and moniliformin. The levels of deoxynivalenol were positively related to F. graminearum levels, as well as to zearalenone levels. Other relationships could not be established. The current approach taken in collecting wheat samples and quantifying the presence of Fusarium Head Blight species and related mycotoxins is an efficient method to obtain insight into the occurrence of these species and toxins in wheat grown under natural environmental conditions. It is recommended that this survey be repeated for several years to establish inter-annual variability in both species composition and mycotoxin occurrence.

Topics: Chromatography, High Pressure Liquid; Crops, Agricultural; Cyclobutanes; Depsipeptides; Food Contamination; Fusarium; Limit of Detection; Mycotoxins; Netherlands; Plant Diseases; Reproducibility of Results; Seeds; Species Specificity; Spectrometry, Mass, Electrospray Ionization; T-2 Toxin; Tandem Mass Spectrometry; Trichothecenes; Triticum; Zearalenone

Mycotoxins such as beauvericin (BEA), deoxynivalenol (DON), enniatin B (ENB), fumonisin B1 (FB1), T-2 toxin and zearalenone (ZEA) can co-occur in food commodities. This aim of this study was to assess the myelotoxicity of these mycotoxins in couple using in vitro human granulo-monocytic (Colony Forming Unit-Granulocyte and Macrophage, CFU-GM) hematopoietic progenitors. Clonogenic assays have been performed in the presence of the following couples of fusariotoxins: DON + BEA, DON + FB1, DON + T-2, DON + ZEA, T-2 + ZEA and BEA + ENB. Co-exposure of human CFU-GM to DON + BEA resulted in synergic myelotoxic effects. The combination of DON + T-2 presented additive or synergic myelotoxic effects. The couples DON + ZEA, T-2 + ZEA and BEA + ENB had additive myelotoxic effects, while the combination of DON + FB1 showed antagonist myelotoxic effects. These in vitro results suggested that the simultaneous presence of mycotoxins in food commodities and diet may be more myelotoxic than the presence of one mycotoxin alone. Diminution of hematopoietic progenitors could give rise to a decrease number of mature blood cells, inducing agranulocytosis and/or thrombocytopenia and in severe cases aplastic anemia.

Topics: Cells, Cultured; Depsipeptides; Fumonisins; Fusarium; Granulocyte-Macrophage Progenitor Cells; Humans; Mycotoxins; T-2 Toxin; Toxicity Tests; Trichothecenes; Zearalenone

Based on a 2-year field trial at two locations in Lower Saxony (Germany), 395 Fusarium isolates belonging to 13 species were collected from more than 3,000 sugar beet roots that were apparently healthy at harvest. In a comparative screen, subsamples were analyzed for Fusarium infection directly after harvest and after different storage conditions. Depending on the storage duration, a different species composition was observed. F. redolens was predominant in freshly harvested beets, while F. culmorum, F. cerealis, and F. graminearum comprised 50.0% (2006) and 84.8% (2007) of the Fusarium mycoflora of sugar beets subjected to long-term pile storage. Randomly selected isolates of all species detected were tested for pathogenicity to sugar beet, but only isolates of F. graminearum and F. sambucinum caused severe root symptoms. Overall, 34 isolates of all species detected were characterized for their mycotoxin profile in rice culture to determine potentially produced toxins for future analysis of sugar beet. A total of 26 Fusarium mycotoxins were detected by liquid chromatography-tandem mass spectrometry, including trichothecenes, zearalenone, and especially high amounts of beauvericin, enniatins, and moniliformin. Further work is required to analyze the natural occurrence of these mycotoxins in sugar beet.

Topics: Beta vulgaris; Cyclobutanes; Depsipeptides; Edible Grain; Fusarium; Germany; Mycotoxins; Oryza; Plant Leaves; Plant Roots; Trichothecenes; Zearalenone

A multi-mycotoxin method was developed for the simultaneous determination of trichothecenes (nivalenol, deoxynivalenol, 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol, neosolaniol, fusarenon-X, diacetoxyscirpenol, HT-2 toxin, T-2 toxin), aflatoxins (aflatoxin-B(1), aflatoxin-B(2), aflatoxin-G(1) and aflatoxin-G(2)), Alternaria toxins (alternariol, alternariol methyl ether and altenuene), fumonisins (fumonisin-B(1), fumonisin-B(2) and fumonisin-B(3)), ochratoxin A, zearalenone, beauvericin and sterigmatocystin in sweet pepper. Sweet pepper was extracted with ethyl acetate/formic acid (99:1, v/v). After splitting up the extract, two-thirds of the extract was cleaned up using an aminopropyl column followed by an octadecyl column. The remaining part was cleaned up using a strong anion-exchange column. After recombination of both cleaned parts of the sample extract, the combined solvents were evaporated and the residue was dissolved in mobile phase; 20 microL was injected into the chromatographic system, so only one run was used to separate and detect the mycotoxins in positive electrospray ionization using selected reaction monitoring. The samples were analyzed with a Micromass Quattro Micro triple quadrupole mass spectrometer (Waters, Milford, MA, USA). The mobile phase consisted of variable mixtures of water and methanol, 1% acetic acid and 5 mM ammonium acetate. The limits of detection of the multi-mycotoxin method varied from 0.32 microg kg(-1) to 42.48 microg kg(-1). The multi-mycotoxin liquid chromatography/tandem mass spectrometry (LC/MS/MS) method fulfilled the method performance criteria required by the Commission Regulation (EC) No 401/2006. Sweet peppers inoculated by Fusarium species were analyzed using the developed method. Beauvericin (9-484 microg kg(-1)) and fumonisins (fumonisin-B(1) up to 4330 microg kg(-1), fumonisin-B(2) up to 4900 microg kg(-1), and fumonisin-B(3) up to 299 microg kg(-1)) were detected.

Topics: Aflatoxins; Capsicum; Chromatography, Liquid; Depsipeptides; Fruit; Fumonisins; Mycotoxins; Ochratoxins; Plant Extracts; Reproducibility of Results; Sterigmatocystin; Tandem Mass Spectrometry; Trichothecenes; Zearalenone

The cytotoxic effects of mycotoxins, induction of reactive oxygen species (ROS) and generation of lipid peroxidation products in CHO-K1 cells were determined as function of increasing time of exposure and concentrations of beauvericin (BEA), patulin (PAT) and zearalenone (ZEA). The end points were evaluated after 24h of exposure, by the tetrazolium salt (MTT) and neutral red (NR) assays. The IC(50) values obtained on the MTT and NR assays ranged from 0.69 to 79.40 microM and 4.40 to 108.76 microM, respectively. To determine the intracellular production of ROS, the intensity of fluorescence emitted from the probe H(2)-DCFDA was measured. The relative intensity of fluorescence from cells incubated with BEA, PAT and ZEA was approximately 4-, 7- and 4-fold higher in comparison with control cells at 0 min, respectively. Lipid peroxidation induced by ROS generation was assessed using the thiobarbituric acid reactive substances (TBARS) method for 2, 24 and 48 h. The malondialdehyde (MDA) production was increased with BEA and PAT exposure in a concentration- and time-dependent manner. MDA was not increased after 1 and 5 microM ZEA exposures for 2h, but was slightly increased at 50 microM. In conclusion, PAT was the most cytotoxic mycotoxin to CHO-K1 cells, followed by BEA and ZEA. Mycotoxins reduce cell viability correlated with the increases of ROS generation and MDA formation in concentration- and time-dependent manner. These data suggested that cytotoxicity and ROS generation are mechanisms of mycotoxins mediated toxicity.

Topics: Animals; CHO Cells; Cricetinae; Cricetulus; Depsipeptides; Lipid Peroxidation; Malondialdehyde; Mycotoxins; Patulin; Reactive Oxygen Species; Zearalenone