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In this study, a dietary exposure assessment of mycotoxins was conducted for the Romanian population using the contamination data of a various categories of wheat-based products for direct human consumption. Wheat-based foods (n = 181) commercialized in Romania, including flour, bread, biscuits, breakfast cereals and pasta, were evaluated by GC-QqQ-MS/MS for the occurrence of deoxynivalenol (DON), 3-acetyldeoxynivalenol (3AcDON), 15-acetyldeoxynivalenol (15AcDON), fusarenon-X, nivalenol, HT-2 and T-2 toxins, diacetoxyscirpenol, neosolaniol and zearalenone (ZEA). DON and 15AcDON were detected in 63 and 5% of all the analyzed samples, whereas 13-AcDON, HT-2, T-2, NIV and ZEA were not detected. Exposure of Romanian adult population was assessed, the EDIs for the sum of DON+3AcDON+15AcDON were 669 ng kg−1 bw day−1 at low-bound estimation, and 690 ng kg−1 bw day−1 at upper-bound estimation, being lower than the TDI set (1000 ng kg−1 bw day−1).

Fusarium sulphureum is a major causal agent of dry rot of potato and fusarium rot of muskmelon fruit. Ozone (O3), a kind of strong oxidant, is widely tested to control postharvest disease in fruit and vegetables by inhibiting the growth of pathogenic fungi. In this study, the effect of ozone treatment on dry rot development, growth of F. sulphureum, diacetoxyscirpenol (DAS) accumulation in vivo and in vitro, and ergosterol biosynthesis were evaluated. The results showed that 2 mg L−1 O3 treatment effectively suppressed dry rot development, DAS accumulation in vivo and in vitro, and significantly inhibited mycelial growth of F. sulphureum. The colony edges and scanning electron microscopy (SEM) observation showed that O3 treatment significantly changed morphological structure of the mycelia; it became shorter, shriveled and rough. Transmission electron microscopy (TEM) results revealed that degradation of organelles intensified, distribution of cytoplasm became uneven, and the integrity of cell membrane was destroyed. Moreover, O3 treatment decreased ergosterol content and DAS accumulation by down-regulating expression of genes involved in ergosterol and DAS biosynthesis pathway. In addition, there was positive correlation between ergosterol content and DAS accumulation in vitro. These results indicate that ozone suppresses dry rot development and DAS accumulation by affecting growth of F. sulphureum and down-regulates expression of genes involved in ergosterol and DAS biosynthesis.

Fusarium Head Blight (FHB) is a destructive disease of small grain cereals and a major food safety concern. Epidemics result in substantial yield losses, reduction in crop quality, and contamination of grains with trichothecenes and other mycotoxins. A number of different fusaria can cause FHB, and there are significant regional differences in the occurrence and prevalence of FHB pathogen species and their associated mycotoxins. Information on FHB pathogen and mycotoxin diversity in Mexico has been extremely limited, but is needed to improve disease and mycotoxin control efforts. To address this, we used a combination of DNA sequence-based methods and in-vitro toxin analyses to characterize FHB isolates collected from symptomatic wheat in Mexico during the 2013 and 2014 growing seasons. Among 116 Fusarium isolates, we identified five species complexes including nine named Fusarium species and 30 isolates representing unnamed or potentially novel species. Significant regional differences (P 0.001) in pathogen composition were observed, with F. boothii accounting for90% of isolates from the Mixteca region in southern Mexico, whereas F. avenaceum and related members of the F. tricinctum species complex (FTSC) accounted for nearly 75% of isolates from the Highlands region in Central Mexico. F. graminearum, which is the dominant FHB pathogen in other parts of North America, was not present among the isolates from Mexico. F. boothii isolates had the 15-acetyldeoxynivalenol toxin type, and some of the minor FHB species produced trichothecenes, such as nivalenol, T-2 toxin and diacetoxyscirpenol. None of the FTSC isolates tested was able to produce trichothecenes, but many produced chlamydosporol and enniatin B.

Fusarium mycotoxins were determined by a multi-mycotoxin liquid chromatography-tandem mass spectrometry (LC-MS/MS) method in cassava ( garri and lafun ) and yam ( amala ) products collected from some selected Nigerian markets. Of the 94 samples analysed, 71% were contaminated with at least one mycotoxin at an incidence rate of 54% ( garri ), 72% ( lafun ), and 82% ( amala ). Deoxynivalenol which ranged between 35 and 99 μg/kg was the most prevalent Fusarium mycotoxin in garri with an incidence rate of 38%, while the dominant mycotoxin in lafun (61%, range = 30–392 μg/kg) and amala (68%, range = 29–155 μg/kg) was fumonisin B 2 . Other mycotoxins detected from the food products included fumonisin B 1 and B 3 , zearalenone, 3-acetyl-deoxynivalenol, 15-acetyl-deoxynivalenol, deoxynivalenol-3-glucoside, T-2 toxin, diacetoxyscirpenol, and fusarenon-X. The consumption of these contaminated products may be a possible route of exposure to a cocktail of Fusarium mycotoxins and can cause potential synergistic health effects on the Nigerian population.

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-17-36 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.

Maize is the staple food of Zimbabweans and is consumed daily in the majority of households, particularly subsistence farming households. The objectives of this study were first, to determine the occurrence of mycotoxins in maize produced and consumed by subsistence farmers in Zimbabwe and second, to determine mycotoxin exposure through maize consumption and subsequently the human health risk. A total of 95 maize meal samples were collected from the household stores of randomly selected subsistence farming households. Maize intake data and agronomic practices of these households were investigated A multi-mycotoxin LC-MS/MS method was used to analyze and quantify mycotoxin contamination in the maize samples. Mycotoxin contamination was compared across agro-ecological zones in order to determine differences in mycotoxin contamination levels and presented. Of the toxicologically relevant mycotoxins, aflatoxin B1 (AFB1), fumonisin B1 (FB1), FB2, deoxynivalenol (DON) and zearalenone (ZEN) were detected in 1, 95, 31, 24 and 15 % of the samples at mean levels of 11, 242, 120, 217 and 110 μg/kg respectively. Other mycotoxins detected in the maize were 15-acetyl-deoxynivalenol (15-ADON), nivalenol (NIV), FB3, alternariol-methylether (AME), AFB2, AFG1 and diacetoxyscirpenol (DAS) and the percentage contamination ranged between 1 and 4 % in the maize samples. Contamination of the maize by the mycotoxins was observed at minimum levels below limit of detection for each mycotoxin and maximum levels of 105, 530, 67, 108, 3, 4 and 14 μg/kg for 15-ADON, NIV, FB3, AME, AFB2, AFG1 and DAS respectively. The median levels of each mycotoxin were reported below the limit of detection, with the exception of FB1 (median, 146 μg/kg), which was further considered in the exposure and risk assessment. Dietary exposure was derived from combining mean maize intake data and median FB1 contamination. Mean maize intake was estimated to be 26.8, 37.2, 30.1, 15.8 and 15.0 g/kg bodyweight (bw)/day for under 5s, children, adolescents, adults and the elderly respectively. Subsequently FB1 exposure from maize was calculated, to be 3.91, 5.40, 4.40, 2.30 and 2.20 μg/kg bw/day for these populations. Exposure to FB1 through maize intake was observed to equate to 196, 272, 220, 115 and 110 % of the provisional maximum tolerable daily intake (2 μg/kg bw/day) for under 5s, children, adolescents, adults and the elderly respectively. Subsistence farming communities in Zimbabwe are at risk of high exposure to FB1 and the risk was highest for under 5s, children and adolescents respectively.

Trichothecenes are groups of chemically related toxins formed by some filamentous fungal species such as Fusarium, Myrothecium, and Stachybotrys. T-2 toxin (T-2), HT-2 toxin (HT-2), neosolaniol (NEO), diacetoxyscirpenol (DAS), monoacetoxyscirpenol (MAS), Verrucarol (VER), Scirpentriol (SCP), and their derivatives are reported as representative type A trichothecenes. Recent advances in the toxicological effects and occurrence of type A trichothecenes as well as their detection and decontamination methods were reviewed in this study. Type A trichothecenes have been detected in numerous food and feed materials and the most popular detection technique was liquid chromatography-tandem mass spectrometry. T-2 and HT-2 were considered as the most prevalent type A trichothecenes in cereals, especially in barley, oats, and animal feeds. Although the concentrations of the detected type A trichothecenes were generally low, severely contaminated samples were observed occasionally which may harm the health of human and animals. Literatures on the occurrence of NEO, MAS, VER, and SCP are still limited. More research on monitoring their prevalence in agricultural products and efficient techniques to reduce their prevalence and harmful effects are needed.

An LC-MS/MS (QqQ) method has been developed and validated for simultaneous determination of the following trichothecenes in UHT cow milk: nivalenol (NIV), deoxynivalenol (DON), deepoxy-deoxynivalenol (DOM-1), 3-acetyldeoxynivalenol (3-ADON), 15-acetyldeoxynivalenol (15-ADON), neosolaniol (NEO), diacetoxyscirpenol (DAS), fusarenon X (FUS-X), T-2 and HT-2 toxins. Sample treatment is simple and based on the extraction with acetonitrile (ACN), acidified with 0.2% formic acid, followed by a purification process, adding sodium acetate to the ACN/water extract in order to separate aqueous phase and, consequently, polar components of the milk. Validation of the method for all the 10 mycotoxins was successful; validation parameters taken into account were as follows: limits of detection (LOD) and quantification (LOQ), linearity, precision (within-day and between-day variability), recovery, matrix effect and stability. The LODs were 10.1, 2.5, 1.5, 1.9, 0.1, 0.5, 1.0, 0.08, 0.4 and 0.05ng/mL for NIV, DON, DOM-1, FUS-X, NEO, 3-ADON, 15-ADON, DAS, HT-2 and T-2, respectively. Mean recovery values (obtained in intermediate precision conditions) were between 63.5 and 75.8 (RSDR≤15%) for all the mycotoxins. All the mycotoxins suffered from matrix effects, especially DON.

Mycotoxins are secondary metabolites produced by filamentous fungi that usually contaminate food products. Coffee is a natural product susceptible to mycotoxin contamination. The present study evaluates the presence of nivalenol, deoxynivalenol, T-2 and HT-2 Toxin, diacetoxyscirpenol, aflatoxin B 1 , aflatoxin B 2 , aflatoxin G 1 , aflatoxin G 2 , fumonisin B 1 , fumonisin B 2 , ochratoxin A, zearalenone, enniatin A, enniatin A 1 , enniatin B, enniatin B 1 , and beauvericin in coffee samples, using liquid chromatography tandem mass spectrometry (LC-MS/MS). The results show that zearalenone was not present in any sample. In the positive samples the contents of fumonisins ranged from 58.62 to 537.45 μg/kg, emerging mycotoxins ranged from 0.10 to 3569.92 μg/kg, aflatoxins ranged from 0.25 to 13.12 μg/kg, and trichothecenes, excepting nivalenol, ranged from 5.70 to 325.68 μg/kg. Nivalenol presented the highest concentrations, from 0.40 to 25.86 mg/kg. Ochratoxin A ranged from 1.56 to 32.40 μg/kg, and five samples exceeded the maximum limit established by the European Commission.

We report the methodology for the quantification of 19 mycotoxins in human plasma using high performance liquid chromatography-mass spectrometry (triple quadrupole). The studied mycotoxins were: deepoxy-deoxynivalenol, aflatoxins (B1, B2, G1, G2 and M1), T-2 and HT-2, ochratoxins A and B, zearalenone, sterigmatocystin, nivalenol, deoxynivalenol, 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol, neosolaniol, diacetoxyscirpenol and fusarenon-X. Sample deproteinization and cleanup were performed in one step using Captiva EMR-lipid (3 mL) cartridges and acetonitrile (with 1% formic acid). The extraction step was simple and fast. Validation was based on the evaluation of limits of detection (LOD) and quantification, linearity, precision, recovery, matrix effect, and stability. LOD values ranged from 0.04 ng/mL for aflatoxin B1 to 2.7 ng/mL for HT-2, except for nivalenol, which was 9.1 ng/mL. Recovery was obtained in intermediate precision conditions and at three concentration levels. Mean values ranged from 68.8% for sterigmatocystin to 97.6% for diacetoxyscirpenol (RDS ≤ 15% for all the mycotoxins). Matrix effects (assessed at three concentration levels and in intermediate conditions) were not significant for most of the mycotoxins and were between 75.4% for sterigmatocystin and 109.3% for ochratoxin B (RDS ≤ 15% for all the mycotoxins). This methodology will be useful in human biomonitoring studies of mycotoxins for its reliability.

A new method for the simultaneous detection of 21 mycotoxins (ochratoxin A, aflatoxin B1, aflatoxin B2, aflatoxin G1, aflatoxin G2, sterigmatocystin, nivalenol, deoxynivalenol, 3-acetyl deoxynivalenol, 15-acetyl deoxynivalenol, diacetoxyscirpenol, neosolaniol, HT-2 toxin, T-2 toxin, fumonisin B1, fumonisin B2, enniatin A, enniatin A1, enniatin B, enniatin B1, and beauvericin) in coffee beverages was internally validated. The method is based on liquid/liquid extraction with a mixture of ethyl acetate/formic acid (95:5 v/v) and detection using triple quadrupole (QqQ) and ion trap (IT) liquid chromatography tandem mass spectrometry. The limits of detection and quantification were 0.02 to 39.64 μg/kg, respectively, and the correlation coefficients were optimal for all mycotoxins (R2 ≥ 0.992). The recovery values ranged from 72% to 97%. The developed method was demonstrated in six real samples of roasted and instant coffee, caffeinated and decaffeinated coffee, and coffee with sugar added. The analyses indicate the presence of the studied mycotoxins in coffee beverages at μg/kg concentrations. Ochratoxin A, a mycotoxin that is regulated in coffee, was detected in two samples under the maximum limit established by a European legislation (CE1881/2006).

In this study a total of 75 commercially Italian samples of baby foods, including 13 infant formula milks (infant formula powders, ready-to-use preparation), 11 dairy products (cheese and yogurt), 25 cereal-based baby foods, 16 fruit and vegetables compotes, and 10 fruit and vegetables purees (composed of pear, peach, banana and for apple), were analyzed to provide an overview on mycotoxin presence. The presence was carried out by evaluating of 23 mycotoxins: ochratoxin-A (OTA), patulin (PAT), two aflatoxins (AFM1, AFB1), three zearalenones (ZONs), which include zearalenone (ZON) and its metabolites (α-zearalenol (α-ZOL) and β-zearalenol (β-ZOL)), nine trichothecenes: deoxynivalenol (DON), 3-and 15-acetyl-deoxynivalenol (3-ADON and 15-ADON), diacetoxyscirpenol (DAS), nivalenol (NIV), fusarenon-X (FUS-X), T-2 toxin (T-2) and HT-2 toxin (HT-2), and five emerging mycotoxins: four enniatins ENs (ENA, ENA1, ENB and ENB1), and beauvericin (BEA). Analytical results showed that 31% of cereal-based baby foods presented levels of mycotoxins. The frequencies of detected mycotoxins, OTA, DON, HT-2, FUS-X, NIV, β-ZOL, ENB, ENB1, ENB4, ENA1 and, BEA, in analyzed samples were 20%, 21%, 3%, 24%, 4%, 7%, 15%, 1%, 5%, 4%, and 1%, respectively. The high value registered was for a cereal-based baby food which contained 832 μg/kg of ENB. 27% of analyzed samples presented co-occurrence of different mycotoxins.

A total of 9 traditional maize based opaque beers were randomly collected from tribal (chewa) rituals and commercial village brewers from Lilongwe and Dowa districts, Malawi in August 2012 for evaluation of mycotoxins using a multi-mycotoxin method based on liquid chromatography-tandem mass spectrometry. Samples were analyzed for aflatoxins B1 (AFB1), AFB2, AFG1, AFG2, fumonisin B1 (FB1), FB2, FB3, zearalenone, ochratoxin A, nivalenol, deoxynivalenol, 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol, neosolaniol, fusarenon-X diacetoxyscirpenol, HT-2 toxin, T-2 toxin, alternariol, alternariol methyl ether, and sterigmatocystin. All beer samples were positive for fumonisins with FB1 + FB2 and FB1 + FB2 + FB3 content of 1745 ± 1294 μg/kg (mean ± std. dev.) and 1898 ± 1405 μg/kg respectively. With exception of one beer sample, all the beers also contained aflatoxins at a mean concentration of 90 ± 95 μg/kg. Consumption of 1.0–6.0 L of the traditional beer from this study translates to daily FB1 + FB2 exposure of 29–174 μg/kg body weight (bw)/day (i.e. >provisional maximum tolerable daily intake of 2 μg/kg bw/day set by the Joint FAO/WHO Expert Committee on Food Additives) and aflatoxin exposure of 1.5–9.0 μg/kg bw/day for a 60 kg adult. The study demonstrated that the consumption of the maize based traditional home-made beer in Malawi can significantly enhance aflatoxins and fumonisin dietary exposure among beer consumers.

A total of 98 samples of oat grain and products from two farming systems were collected during the period of 2009–2011 and analyzed for the presence of trichothecenes (deoxynivalenol, nivalenol, 3-acetyldeoxynivalenol, T-2 toxin, HT-2 toxin, monoacetoxyscirpenol, diacetoxyscirpenol) and zearalenone using liquid chromatography–tandem mass spectrometry method. All mycotoxins were found at different frequency and levels in the tested oat samples; however, a higher intensity of distribution of DON, T-2 and HT-2 toxins was found in the conventional system of farming than in organic. The highest concentration in both systems (up to 655 μg kg−1) was that of the mycotoxin NIV.

A new analytical method for the rapid and simultaneous determination of ten mycotoxins including patulin, zearalenone and eight trichothecenes (nivalenol, fusarenon-X, diacetoxyscirpenol, 3-acetyl-deoxynivalenol, neosolaniol, deoxynivalenol, T-2 and HT-2) in wheat semolina has been developed and optimized. Sample extraction and purification were performed with a modified QuEChERS-based (acronym of Quick, Easy, Cheap, Effective, Rugged and Safe) procedure and determined by gas chromatography (GC) coupled to triple quadrupole instrument (QqQ). This is the first paper on the application of GC-QqQ-MS/MS to analysis of mycotoxins. Careful optimization of the gas chromatography-tandem mass spectrometry parameters was achieved in order to attain a fast separation with the best sensitivity allowing a total run time of 16 min. The validation was performed by analyzing recovery samples at three different spiked concentrations, 20, 40 and 80 μg kg(-1), with four replicates (n=4) at each concentration. Recoveries ranged from 74% to 124% and the intra-day precision and inter-day precision, expressed as relative standard deviation, were lower than 13% and 17%, respectively for all studied compounds, except for zearalenone. Limits of quantification (LOQ) were lower than 10 μg kg(-1) for all studied mycotoxins. Eight concentration levels were used for constructing the calibration curves which showed good linearity between LOQ and 100 times LOQ concentration levels (linear range). Matrix-matched calibration for applying the method in routine analysis is recommended for reliable quantitative results. The method validated was successfully applied to fifteen wheat semolina samples detecting occurrence of mycotoxins at concentrations below the maximum permissible level.

A previous developed matrix solid-phase dispersion (MSPD) extraction method was applied for the routine analysis of aflatoxins (AFs), ochratoxin A (OTA), fumonisins (FB 1 and FB 2 ), beauvericin (BEA), nivalenol (NIV), deoxynivalenol (DON), the toxin T-2 (T-2), toxin HT-2 (HT-2), diacetoxyscirpenol (DAS) and zearalenone (ZEN) in tiger-nuts by liquid chromatography–triple-quadrupole linear ion trap (HPLC–QTRAP ® ). The extraction solid support used was C 18 , while the elution solvent was acetonitrile/methanol (50/50, v/v) 1 mM ammonium formate. Using matrix-matched calibration, recoveries and repeatabilities were in the range 67–89% and 2–11% relative standard deviation (RSD), respectively. The method was applied to determine the occurrence of the fourteen selected mycotoxins in a total of 83 tiger-nut samples purchased from different local markets of Valencian Community (Spain) during (March–June 2010 and March–May 2011). DON, OTA, AFs and BEA were detected in 26 samples of the total number of samples.

Malting and brewing by-products (germ/rootlets, spent grains and spent yeast) represent a valuable and nutritious source of food for the livestock. The incidence of mycotoxins in animal feed is not an unusual phenomenon, and for that reason, feed obtained from malting and brewing process is subjected to mycotoxicological anyalysis. Objectives of this study were to establish the impact of the initial F. culmorum contamination and fungicide treatment on trichothecenes, namely deoxynivalenol (DON), 3-acetyl deoxynivelanol (3- ADON), nivalenol (NIV), diacetoxyscirpenol (DAS), fusarenon X (FUS X), HT-2 and T-2 . Their concentrations were measured in water after steeping, germ/rootlets, spent grains and spent yeast. Two wheat genotypes (A and B) were treated in four different ways in the field: control (a), treated with fungicide (b), inoculated with F. culmorum spores and treated with fungicide (c), and inoculated with F. culmorum spores (d). Standard malting procedure for wheat malt was implemented, followed by degermination process and brewing procedure. Mycotoxins were determined using multi- mycotoxin LC-MS/MS method. DON proved to be the most occurring mycotoxin in all of the samples and the highest concentration was found in steeping water of sample A(d) where detected concentration of DON was 20326 µgL-1. Germ/rootlets contained DON, 1384 μg/L in B(d) and 5636 μg/L in A(d) sample. Spent grains showed elevated DON value (1178 μg/L) for B(b) sample. Spent yeast showed increased DON values for A(d) sample (705 μg/L) but also for B(d) sample (1180 μg/L).

The toxin producing capacity of seven Fusarium species (F. langsethiae, F. sporotrichioides, F. poae, F. avenaceum, F. tricinctum, F. graminearum and F. culmorum) and the effect of culture conditions on the toxin production were studied. The strains were isolated from Finnish grains and cultivated on a grain mixture at three different water activity/temperature combinations (i.e. 0.994/15 degrees C; 0.994/25 degrees C; 0.960/25 degrees C). The mycotoxins produced were analyzed with a multi-toxin method based on liquid chromatography-tandem mass spectrometry enabling the simultaneous determination of 18 different Fusarium toxins. The general toxin profiles revealed F. langsethiae and F. sporotrichioides as producers of diacetoxyscirpenol, neosolaniol, HT-2 and T-2-toxins. F. sporotrichioides produced additionally beauvericin. In the F. poae cultures, only beauvericin was detected. F. avenaceum and F. tricinctum were capable of producing enniatins, moniliformin and antibiotic Y, and F. graminearum and F. culmorum produced zearalenone, deoxynivalenol and 3-acetyl deoxynivalenol. Differences existed in the quantitative toxin production between the individual strains representing the same species. Additionally, the culture conditions affected the range and amounts of toxins produced. In general, a(w) 0.994 and temperature of 15 degrees C favoured the type-A trichothecene production of F. langsethiae and F. sporotrichioides. The beauvericin production of F. sporotrichioides occurred more favourably at a(w) 0.960 and 25 degrees C. F. poae produced the highest concentrations of beauvericin under two different conditions, namely at a(w) 0.994/15 degrees C and a(w) 0.960/25 degrees C. None of the combinations particularly favoured toxin production of F. avenaceum, with all three toxins being produced extensively at all culture conditions. F. tricinctum produced enniatins most efficiently at a(w) 0.994/25 degrees C. The moniliformin production of both these two species occurred readily at a(w) 0.960/25 degrees C. F. culmorum and F. graminearum produced the highest concentrations and variety of mycotoxins at a(w) 0.960/25 degrees C. The results give valuable information on the toxigenicity of some important Fusarium species. Additionally, this is the first in-depth study to investigate the influence of environmental conditions on the toxin production by F. langsethiae, F. poae, F. avenaceum and F. tricinctum.

A suitable extraction and purification method for the simultaneous liquid chromatography-mass spectrometry (LC-MS) determination of five mycotoxins, three type A, diacetoxyscirpenol (DAS), T-2 toxin (T-2) and HT-2 toxin (HT-2), and two type B-trichothecenes, deoxynivalenol (DON) and nivalenol (NIV), has been optimised using a modified "Quick Easy Cheap Effective Rugged and Safe" (QuEChERS) method. Different solvents were studied in the extraction procedure to obtain better recoveries, which ranged from 86 to 108%, using a 85/15 (v/v) mixture of methanol/acetonitrile. The values obtained for recovery, repeatability and reproducibility of the optimized method are in agreement with Commission Directive 2005/26/EC for methods of analysis of Fusarium toxins. Finally, this optimized procedure was applied in wheat flour samples commercialized in Spain.

In this study, the direct detection of toxigenic fungi and the quantification of trichothecene levels in food and feed materials from India were investigated. A total of 40 different food and feed materials were subjected to PCR for the detection of toxigenic Fusarium spp. PCR indicated the presence of tri5 gene in seven samples (six sorghum samples and one poultry feed). Results indicated the presence of T-2 toxin and diacetoxyscirpenol (DAS) in all the six sorghum samples whereas only T-2 content was noticed in poultry feed. Levels of T-2 and DAS varied from 0.012 (±0.004) to 0.13 (±0.03) mg/kg and 0.014 (±0.004) to 0.084 (±0.004) mg/kg, respectively. The levels of these toxins were falling within the range detected previously in India and in other parts of the world.

Fusarium species are among the most studied plant-pathogenic fungi, with several species causing diseases on maize, wheat, barley, and other food and feed grains. Decreased yield, as well as diminished quality and value of the grain, results in significant worldwide economic losses. Additionally, Fusarium species produce a chemically diverse array of mycotoxins such as diacetoxyscirpenol, deoxynivalenol, nivalenol, T-2 toxin, zearalenone, fumonisins, fusarin C, beauvericin, moniliformin, and fusaproliferin. The dominant Fusarium species associated with feed grain that produce these mycotoxins are reviewed with emphasis on their current taxonomy, phylogenetic relationships, and general biology. Ecological and environmental factors associated with plant-fungal interactions and potential mycotoxin contamination of feed also are discussed with primary emphasis on two main diseases: head blight of small grains and ear rot of maize. The past quarter-century has provided much detail on the morphology, physiology, genetics and genomics of Fusarium species. Such data are critical for understanding these fungi and for managing their impact on the safety, value, and yield of quality grain.

The purpose of this study was to investigate the diacetoxyscirpenol (DAS, anguidine) level of contaminated cereal and pulse products in Turkey. DAS was detected using high performance liquid chromatography (HPLC) with diode array detector (DAD) at 205 nm and suspicious the results for two specimens suspected to contain DAS were confirmed by gas chromatography–mass spectrometry (GC–MS). An acetonitrile–water (21 + 4, v/v) extract of the sample was cleaned up on a column packed with alumina/charcoal (0.35 g + 0.40 g). The minimum detectable amount of DAS was 16 ng/injection, limit of detection was 0.8 μg/g for HPLC. A total of 85 commercially available cereal and pulse product samples, collected from markets and street bazaars, were analyzed. The recoveries for corn flour with the known amounts of DAS (2, 3, 4 μg/g) were 85.3% (SD 4.81, n = 5), 98.1% (SD 12.6, n = 5) and 96.4% (SD 3.2, n = 5), respectively. DAS was detected in none of the cereal and pulse products.

The degradation of 12 trichothecene mycotoxins by chicken intestinal microbes was monitored by liquid chromatography-ultraviolet-mass spectrometry under positive ion atmospheric pressure chemical ionization. Two pathways were observed: deacylation and deepoxidation. Essentially complete conversions to the deepoxy metabolites were observed for the non-acylated trichothecenes 4-deoxynivalenol, nivalenol, and verrucarol. However, deacetylation was the predominant pathway for the monoacetyl trichothecenes 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol (15ADON), and fusarenon X. Small amounts of the deepoxy metabolites were observed from 15ADON and large amounts from 15-monoacetoxyscirpenol where steric hindrance protected the C-15 acetyl groups from enzymatic attack. Diacetylated trichothecenes diacetoxyscirpenol and neosolaniol exhibited only deacetylation. The larger isovaleryl functionality was resistant to removal and deepoxidation was the prevalent reaction in HT-2 toxin and T-2 triol, whereas T2 toxin showed only deacetylation.

Trichothecene mycotoxins are common contaminants of cereal grains and animal feed worldwide. The toxins are toxic to both human and animals. The objectives of this study were to determine the occurrence of trichothecenes in grains and animal feed in Croatia. Total of 465 samples were collected during the seven-year period (1998–2004) from manufactures and small holders farm storage facilities. The samples were analyzed by thin layer chromatography, which proved to be fast, reliable and inexpensive method. T-2 toxin, diacetoxyscirpenol and deoxynivalenol were detected in 16.8%, 27.6% and 41.2%, respectively. The amount of toxins ranged between 0.05 and 3.4 mg/kg. The majority of animal feed samples was poultry feed. Only small number of it contained T-2 toxin and diacetoxyscirpenol levels greater than the Croatian regulatory levels for poultry feed. Positive samples were in correlation with evidenced clinical symptoms of toxicosis in poultry. Since trichothecenes are frequently isolated from animal feed and grains in Croatia, they could have significant economic and safety implications in animal production.

Several studies have shown that the mycotoxins T-2 toxin, diacetoxyscirpenol (DAS), deoxynivalenol (DON) and nivalenol (NIV) affect lymphocyte functioning. However, the molecular mechanisms underlying the immunomodulatory effects of these trichothecenes are not defined yet. In this study, the potency of the type A trichothecenes T-2 toxin and DAS, and the type B trichothecenes DON (and its metabolite de-epoxy-deoxynivalenol; DOM-1) and NIV to reduce mitochondrial activity and to induce apoptosis of Jurkat T cells (human T lymphocytes) were examined. T-2 toxin and DAS are much more cytotoxic at low concentrations than DON and NIV as shown by the AlamarBlue cytotoxicity assay. In addition, the mechanism whereby DON and NIV induced cytotoxicity is mainly via apoptosis as we observed phosphatidylserine externalization, mitochondrial release of cytochrome c, procaspase-3 degradation and Bcl-2 degradation. In contrast, type A trichothecenes reduce the mitochondrial activity at approximately 1000-fold lower concentrations than the type B trichothecenes, resulting in necrosis. These data suggest that the mechanisms resulting in cytotoxic effects are different for type A and type B trichothecenes.

The trichothecenes deoxynivalenol (DON), nivalenol (NIV), diacetoxyscirpenol (DAS), and toxins T2 (T2) and HT2 (HT2) were searched for in 78 samples of corn-based products sold in the city of Sao Paulo/Brazil. Acetonitrile:water was used for extraction and the clean-up was accomplished with Mycosep columns. Trifluoroacylation was employed before quantification and confirmation by gas chromatograph with a selective mass detector (ion trap) in electron ionization mode. The detection limits varied with the several toxin/matrix combinations and ranged from 20 to 120 ng/g. The recoveries ranged from 71 to 155%. Only two samples were found contaminated: a sample of corn grits with 555 ng/g of T2 and 767 ng/g of HT2 and a sample of corn flour that exhibited traces of DON and NIV. DAS was not detected in any sample. The results indicate a low occurrence of trichothecene mycotoxins in corn-based products commercialized in the city of Sao Paulo in spite of high levels of T2 and HT2 found in one sample and show no immediate cause of concern. Nonetheless more extensive surveys conducted for several years are advisable in order to furnish a more complete picture of the incidence of these toxins in Brazilian corn products.

Landraces of maize ( Zea mays ssp. mays ) and its wild teosinte relatives ( Zea mays spp. parviglumis and mexicana ) were surveyed for sensitivity to fumonisin B 1 , a phytotoxin produced by the maize pathogen Gibberella moniliformis. Only two of 42 Z. mays samples were highly insensitive to FB 1 (ED 50 = ca. 200 μM). The teosintes and 76% of the maize landraces were moderately or highly sensitive to FB 1 (ED 50 ⩽ 30 μM), which indicates that FB 1 sensitivity is likely to be an ancestral trait in Z. mays . F 1 generations derived from crosses between FB 1 -sensitive maize inbred B73 and insensitive landraces were significantly less sensitive than B73. Thus, our data indicate that FB 1 -insensitivity is a relatively rare but heritable trait in maize. We also report the sensitivity of maize to other Gibberella toxins – beauvericin, diacetoxyscirpenol, and moniliformin.

A total of 219 samples of foodstuffs of plant origin, consisting of grain-based food, pseudocereals and gluten-free food as well as vegetables, fruits, oilseeds and nuts, were randomly collected during 2000 and 2001 in food and health food stores. A spectra of 13 trichothecene toxins including diacetoxyscirpenol (DAS), 15-monoacetoxyscirpenol (MAS), scirpentriol (SCIRP), T-2 and HT-2 toxins (T-2, HT-2), T-2 triol, T-2 tetraol, neosolaniol (NEO) of the A-type as well as deoxynivalenol (DON), 3- and 15-acetyl-DON (3-, 15-ADON), nivalenol (NIV), and fusarenon-X (FUS-X) of the B-type were determined by gas chromatography/mass spectrometry. Analysis of zearalenone (ZEA), alpha- and beta-zearalenol (alpha- and beta-ZOL) was made by high-performance liquid chromatography with fluorescence and UV-detection. Detection limits ranged between 1 and 19 microg/kg. Out of 84 samples of cereal-based including gluten-free foods, 60 samples were positive for at least one of the toxins DON, 15-ADON, 3-ADON, NIV, T-2, HT-2, T-2 tetraol and ZEA, with incidences at 57%, 13%, 1%, 10%, 12%, 37%, 4% and 38%, respectively, whereas SCIRP and its derivatives MAS and DAS, T-2 triol, Fus-X as well as alpha- and beta-ZOL were not detected in any sample of this subgroup. Contents of DON ranged between 8 and 389 microg/kg, for all other toxins determined concentrations were below 100 microg/kg. The pseudocereals amaranth, quinoa and buckwheat were free of the toxins investigated. Ten of 85 samples of vegetables and fruits were toxin positive. ZEA and the type A trichothecenes MAS, SCIRP, DAS, HT-2 were detected in 7, 3, 2, 1 and 1 samples, respectively. Out of 35 samples of oilseeds and nuts, 7 samples were toxin positive. HT-2, T-2 and ZEA were detected in 4, 3 and 4 samples, respectively. In vegetables and fruits as well as in oilseeds and nuts, toxin levels were below 50 microg/kg. None of the B-type trichothecenes analysed was found for both subgroups.

In 2001 the SCOOP (SCOOP: Scientific Co-operation on Questions relating to Food) task 3.2.10 "Collection of occurrence data of Fusarium toxins in food and assessment of dietary intake by the population of EU Member States" was established. The task was divided in three subtasks (zearalenone, fumonisins and trichothecenes). Results of the subtask trichothecenes, which is co-ordinated by The Netherlands, will be presented. About 35,000 results were received about occurrence of 12 different trichothecenes (deoxynivalenol (DON), nivalenol (NIV), 3 and 15 acetyl-deoxynivalenol (3/15-AcDON), fusarenon-X (FUS-X), T-2 and HT-2 toxin, T2-triol, diacetoxyscirpenol (DAS), neosolaniol (NEOSOL, monoacetoxyscirpenol (MAS) and verrucarol (VOL)) in various food and food raw materials from 12 countries. Only the results of DON, NIV, T-2 and HT-2 toxin are included in this paper, because most of the data refer to these toxins and only for these trichothecenes the Scientific Committee for Food sets (temporary)-Tolerable Daily Intakes (t-TDIs). Occurrence data: By far most of the occurrence data were obtained for DON in wheat. Among cereals, corn showed the highest level of contamination with trichothecenes. Consumption data: There is a significant lack of consumption data in some countries. In particular information on baby's and children's food is generally not available. Intake data: Wheat and wheat containing products (like bread and pasta) represent the major source of intake for the four trichothecenes. The mean intakes for DON are below the TDI, however for the young children groups the mean intakes are sometimes (very) close to the TDI. By comparing the high intake levels for DON with the TDI, it is clear that especially for the young children groups most of the intakes are above the TDI. For NIV, the (mean and high level) intakes are far below the TDI. The summarised T-2 and HT-2 toxin intakes are in most cases (for the mean as well as the high level intake) above the t-TDI.

In 1999 and 2000, high concentrations of deoxynivalenol (DON) were found in wheat and breakfast cereals sold on the Dutch market. Besides wheat other grains can be contaminated with trichothecenes. Using these grains in the brewing process, the produced beer could be contaminated with trichothecenes. A previously developed GC–FID method to determine trichothecenes in wheat was made applicable for the determination of trichothecenes (DON, nivalenol, 3-acetyldeoxynivalenol, fusarenon X, T-2 toxin, HT-2 toxin, diacetoxyscirpenol and neosolaniol) in beer. Surveys were carried out in November 2000 and March 2001 to investigate the presence of trichothecenes in beers marketed in the Netherlands. In only three of the 51 beer samples low quantities of DON were found, ranging from 26 to 41 μg/l. Besides DON, no trichothecenes were found in the beers at levels above the limit of quantification (25 μg/l). All positive samples were from so-called top fermentation beers.

A total of 843 commercial animal feed and foodstuff samples (465 samples of agricultural commodities and 378 samples of animal feeds) from all over the Kingdom of Saudi Arabia were collected during the years 1997 to 2000 and analyzed for type A and type B trichothecenes (diacetoxyscirpenol, neosolaniol, HT-2 toxin, T-2 toxin, nivalenol, fusarenon-x, deoxynivalenol). Levels of mycotoxins detected ranged from2 to 4,000 microg/kg deoxynivalenol, 3.25 to 500 microg/kg fusarenon-x, 3.13 to 600 microg/kg nivalenol, 3.13 to 50 microg/kg diacetoxyscirpenol, 6.25 to 200 microg/kg neosolaniol, 3.13 to 18.75 microg/kg HT-2 toxin, and 6.25 microg/kg T-2 toxin. The study reflected the need for routine surveillance of agricultural commodities to minimize potential hazards to human health.

Trichothecenes are mycotoxins produced by several fungal genera. The Fusarium species, mainly, can contaminate a wide range of cereals used for human and animal consumption. Trichothecenes are associated with various adverse health effects in animals and humans. Deoxynivalenol (DON) and nivalenol (NIV) are the trichothecenes most commonly found worldwide, although 3-acetyldeoxynivalenol, fusarenon X, T-2 toxin, HT-2 toxin, diacetoxyscirpenol (DAS) and neosolaniol are also found. They are included in the present study. For the determination of these trichothecenes in wheat, a method based on capillary gas chromatography (GC) with flame ionisation detection (FID) has been developed and validated. The trichothecenes are extracted from the sample matrix by acetonitrile/water (84/16, v/v). Two different Mycosep® clean-up columns are used to purify the extract. The extract is evaporated to dryness and the trichothecenes are derivatised to trimethylsilyl ethers at room temperature. The residue is dissolved in iso-octane and washed with water. The final extract is analysed for trichothecenes by GC with FID. Quantification is based on the internal standard α-chloralose. The average recoveries for the trichothecenes range from 79% for NIV to 116% for DAS. The limit of quantification is 75 μg/kg for each of the individual trichothecenes. The GC-FID method produced good results in an intercomparison study of trichothecene analysis within the European Union Standards, Measurements and Testing Programme. A survey was carried out in the Netherlands in 1999 to detect the presence of trichothecenes in imported wheat. A temporary tolerance limit of 500 μg/kg is in effect in the Netherlands for DON in cleaned wheat. Seven of the 22 wheat samples exceeded this limit; one exceeded the limit by more than 100%. Thirteen of the 22 wheat samples exceeded a proposed DON tolerance limit of 120 μg/kg for cleaned wheat. Indeed, 12 samples exceeded the limit by more than 100%. Besides DON, no trichothecenes were found in the wheat samples at levels above the limit of quantification.

The effects of feeding T-2 toxin, diacetoxyscirpenol (DAS), or aflatoxin B1 at levels up to 1,000 ppb for 5 weeks on performance, health, and immune response of enterally and parenterally immunized chicks were examined. No decreases in growth or feed efficiency were observed when T-2, DAS, or a mixture of these mycotoxins were fed for 35 days. Aflatoxin at concentrations above 800 ppb resulted in decreased growth and feed efficiency after 4 weeks. Feeding T-2 and DAS resulted in oral lesions and mild intestinal inflammation, but no other pathological or histopathological lesions. Aflatoxin caused enlargement and discoloration of liver and kidneys and mild intestinal inflammation. No effects of T-2, DAS, or aflatoxin B1 were observed on antibody production to antigens administered by enteral or parenteral routes.

Three experiments were conducted to determine the effects of 4,15-diacetoxyscirpenol (DAS) on BW, feed consumption, and oral lesions of broiler breeders. In Experiment 1, caged broiler breeder hens were fed 0, 5, 10, or 20 mg DAS/kg diet from 24 to 25 wk of age. There were dose-related decreases in BW and feed consumption indicating feed refusal, as well as dose-related increases in the extent of mouth lesions. The areas of the mouth most sensitive to DAS were associated with the salivary glands and the tip of the tongue. In Experiment 2, individually caged male and female broiler breeders were fed a basal diet containing 0, 5, 10, or 20 mg DAS/kg from 25 to 27 wk of age. There were dose-related decreases in BW and feed consumption for the female broiler breeders, whereas there was a decrease in feed consumption for the male broiler breeders at the 10 and 20 mg DAS/kg levels. In Experiment 3, male broiler breeders were fed 0 or 10 mg DAS/kg diet from 23 to 25 wk of age on a litter floor. For this experiment the daily intake of feed was restricted, and the feed consumption rate was measured. There was an increased amount of unconsumed feed at 23 wk of age due to the presence of DAS. In summary, the experiments provided evidence that DAS caused decreased BW and feed consumption as well as cytotoxic injury including oral lesions in broiler breeders.

Two experiments were conducted to determine the effects of 4,15-diacetoxyscirpenol (DAS) on fertility and hatchability of broiler breeders. In Experiment 1, naturally mated broiler breeders were studied. A limited daily allocation of feed containing 0 (basal), 1.25, 2.5, or 5.0 mg DAS/kg diet was provided from 67 to 69 wk of age in slat-litter floor pens. Fertility was consistently improved by the 5.0 mg/kg level of DAS and intermittently by the 1.25 and 2.50 mg/kg levels. The effect disappeared upon removal of DAS. In Experiment 2, individually caged broiler breeder males and females were fed a basal diet containing 0, 5, 10, or 20 mg DAS/kg diet from 25 to 27 wk of age. Semen was pooled from males within each treatment and used to inseminate females from each treatment in a 4 x 4 factorial design. Female-related fertility was increased at the 5 and 10 mg DAS/kg levels and male-related fertility was decreased by the 10 and 20 mg DAS/kg levels. Small, fluid-filled cysts were observed on the testes of many DAS-treated males upon necropsy. In summary, low levels of DAS (or =10 mg DAS/kg) appeared to improve female-related fertility, presumably because of enhanced spermatozoal storage within the oviduct. Conversely, DAS (or =10 mg DAS/kg) decreased male-related fertility, presumably by direct toxic effects on the testes. Overall, it appeared that levels of DAS below 5 mg DAS/kg feed would not be detrimental to fertility and hatchability.

Trichothecenes, zearalenone (ZEN) and fumonisins are the major Fusarium mycotoxins occurring on a worldwide basis in cereal grains, animal feeds and forages. Other important Fusarium mycotoxins include moniliformin and fusaric acid. Spontaneous outbreaks of Fusarium mycotoxicoses have been recorded in Europe, Asia, New Zealand and South America and, in addition, chronic exposure occurs on a regular and more widespread scale. The metabolism and adverse effects of the Fusarium mycotoxins are considered in this review with particular reference to recent data on specific and proposed syndromes and to interactions among co-occurring mycotoxins. Within the trichothecene group, deoxynivalenol (DON) is associated with emesis, feed refusal and depressed feed intake in pigs, while T-2 toxin and diacetoxyscirpenol (DAS) are now clearly linked with oral lesions in poultry. The gut microflora of farm livestock are able to transform DON to a de-epoxy derivative. In contrast, the ovine metabolism of ZEN results in the production of five metabolites and relatively high levels of these forms may be excreted in the urine as glucuronides. There is now undisputed evidence that ZEN and its metabolites possess estrogenic activity in pigs, cattle and sheep, but T-2 toxin has also been implicated in reproductive disorders in farm livestock. Fumonisins are positively linked with pulmonary edema in pigs, leukoencephalomalacia in equines and with deranged sphingolipid metabolism in these animals. Fusarium mycotoxins have also been provisionally implicated in ovine ill-thrift, acute mortality of poultry and in duodenitis/proximal jejunitis of horses. Several Fusarium mycotoxins may co-occur in a particular feed ingredient or in compound feedingstuffs. In general, combinations of Fusarium mycotoxins result in additive effects, but synergistic and/or potentiating interactions have been observed and are of greater concern in livestock health and productivity. Synergistic effects have been reported between DON and fusaric acid; DON and fumonisin B 1 (FB 1 ); and DAS and the Aspergillus -derived aflatoxins. Limited evidence of potentiation between FB 1 and DON or T-2 toxin has also emerged recently. Additive and synergistic effects between known and unidentified mycotoxins may account for enhanced adverse effects observed on feeding Fusarium -contaminated diets. The potential for transmission of DON into eggs and of ZEN into porcine kidney and liver has been demonstrated. However, lactational carry-over of FB 1 appears not to occur, at least in cows and sows. It is concluded that livestock health, welfare and productivity may be severely compromised by consumption of DON, T-2 toxin, DAS, ZEN and fumonisins and by interactions among these mycotoxins. Safety of some animal products may also be at risk. Furthermore, in view of the limited options available for remediation, it is concluded that exploitation of crops resistant to Fusarium infection offers the most viable strategy for reducing mycotoxin contamination of grain and animal feed.

The trichothecenes are mycotoxins produced by fungi of the genus Fusarium, which are commonly present in foods and feed of cereal origin. Owing to the lack of sufficient toxicological data for most of the trichothecenes, in vitro studies may contribute to risk assessments of these toxins. In the present report, human lymphocyte cultures were used to study the individual variation in sensitivity among humans and the effects on in vitro Ig production. Furthermore, proliferative responses of cells exposed to combinations of two of the toxins were studied. Four toxins, T-2 toxin, diacetoxyscirpenol (DAS), nivalenol (NIV) and deoxynivalenol (DON) were included in the study. All four of the tested trichothecenes effectively inhibited mitogen-induced lymphocyte proliferation. There were no statistically significant differences in sensitivity to the toxins between lymphocytes from female and male blood donors. The individual variation in sensitivity, evaluated as the range of IC50 values, was rather limited (within a factor of 3 to 4). Immunoglobulin production by pokeweed-stimulated human lymphocytes was also effectively inhibited with IC50 values similar to the IC50 values in the proliferation tests for DON and NIV. However, IC50 values for Ig synthesis in cultures exposed to T2 were approximately two to three times higher than the corresponding IC50 values found in the proliferation tests. At low levels of exposure, elevated Ig production was observed in lymphocyte cultures from four out of the five blood donors tested. This effect was most pronounced on IgA synthesis. Combinations of NIV with T2, DAS or DON resulted in additive toxicity in the lymphocyte proliferation test, while combinations of DON with T2 or DAS resulted in an inhibition that was slightly lower than what could have been expected from the inhibition produced by the individual toxins. In conclusion, the tested trichothecenes inhibited both proliferation and Ig production in human lymphocytes in a dose-dependent manner with limited variation in sensitivity between individuals. Enhanced Ig production was observed in cell cultures exposed to the lower doses of the toxins. Combined exposure to two of the toxins resulted mainly in additive or antagonistic effects, although synergistic effects cannot be excluded and should be further investigated. These findings indicate that the total intake of type A and B trichothecenes should be taken into account in risk assessments.

An effective method for the determination of seven trichothecenes – deoxynivalenol (DON), nivalenol (NIV), T-2 tetraol (T-24), fusarenon-X (FUS-X), diacetoxyscirpenol (DAS), T-2 toxin (T-2), HT-2 toxin (HT-2) in cereals is presented. Gel permeation chromatography on Bio-Beads S-X3 was used for clean-up of acetonitrile–methanol extract. GC–ECD was used for identification and quantification of trifluoroacetylated trichothecenes. The limit of quantitation for the method was in the range 40–200 μg/kg. Recoveries at a spiking level of 2 mg/kg ranged from 76 to 100%.

Two clean-up principles for the multi-determination of seven trichothecene toxins, deoxynivalenol, nivalenol, diacetoxyscirpenol, fusarenon-X, T-2 tetraol, HT-2 toxin and T-2 toxin in wheat extract are described. For clean-up of acetonitrile–water (84:16, v/v) extract either gel permeation chromatography (Bio-Beads S-X3 gel) or solid-phase extraction (combination of Florisil and C18-silica gel SPE cartridges or a Romer Labs. MycoSep 225 column) were used. The MycoSep 225 column was chosen as the best alternative for clean-up of grain samples. Recovery of this procedure was tested on certified material. Derivatisation of analytes prior to the final determinative step was carried out by trifluoroacetic acid anhydride. Trifluoroacyl derivatives of the trichothecenes were separated by high-resolution capillary gas chromatography with electron-capture detection.

A method for qualitative and quantitative analysis of Fusarium mycotoxins by gas chromatography-mass spectrometry (GC-MS) using cold on-column injection was improved. Eight typical mycotoxins, including deoxynivalenol (DON), 3-acetyldeoxynivalenol (3ADN), fusarenon-X (FX), diacetoxyscirpenol (DAS), 15-monoacetylscirpenol (15MAS), T-2 toxin (T-2), scirpentriol (SCT), and zearalenone (ZEA) were subjected to GC-MS without chemical derivatization by means of the on-column injection technique. Chromatographic separation of the toxins extracted from barley was achieved as a single peak, and the specific EI mass spectra of each toxin were obtained. The fatty acids in the extract that interfere with measurements of the toxins on the gas chromatogram were removed by precipitation as an insoluble metal soap with zinc acetate. Additional clean-up was accomplished using a Bond Elut Florisil cartridge. The quantitative detection limit in barley ranged from 0.1 to 0.5 micrograms/g. The average recoveries of 93.1% for DON, 3ADN, 15MAS, DAS, T-2 and ZEA, and 46.0% for FX and SCT added to barley at the level of 1 microgram/g were obtained.

The individual and combined effects of feeding diets containing 300 mg fumonisin B1 (FB1), and 4 mg diacetoxyscirpenol (DAS) or 3 mg ochratoxin A (OA) were evaluated in two experiments using female turkey poults (Nicholas Large Whites) from day of hatch to 3 wk of age. When compared with controls, body weight gains were reduced 30% (Study 1) and 24% (Study 2) by FB1, 30% by DAS, 8% by OA, 46% by the FB1 and DAS combination, and 37% by the FB1 and OA combination. The efficiency of feed utilization was adversely affected by all treatments except FB1 in Experiment 2. Relative weights of the liver were significantly increased by all treatments except the DAS treatment. Serum concentrations of cholesterol were decreased and activities of aspartate aminotransferase and lactate dehydrogenase were increased and several hematological values were altered in poults fed FB1 alone and in combination with either DAS or OA. Results indicate additive or less than additive toxicity, but not toxic synergy, when poults are fed diets containing 300 mg FB1, and 4 mg DAS or 3 mg OA/kg of diet. The likelihood of encountering FB1, DAS, or OA at these concentrations in finished feed is small. However, under field conditions, other stress factors could alter the impact of these mycotoxins on the health and performance of poultry.

A survey of the natural occurrence of trichothecenes and zearalenone in samples from Buenos Aires and Santa Fe provinces, Argentina, was carried out in six different crops during harvest. Among 1056 samples analysed, 524 (49.62%) were contaminated with trichothecenes and zearalenone. Deoxynivalenol was detected in all of the contaminated samples, 3-Acetyl DON in 22 samples and zearalenone in 78. In 1986, T-2 toxin (20 samples), HT-2 toxin (18), neosolaniol (16) and diacetoxyscirpenol (26) were also found. The results suggest that when meteorological conditions favour contamination it is necessary to carry out strict controls on harvest time.

A yeast (Kluyveromyces marxianus) bioassay was used to establish the relative toxicity of 16 trichothecenes and some of their interactions. Removal of an isovaleryl group, one or two acetyl groups, or two acetyl plus an isovaleryl groups from T-2 toxin (T2) to form diacetoxyscirpenol, HT-2 toxin (HT2), T2-triol and T-2 tetraol (T2-4ol) decreased toxicity 7, 36, 276 and 558-fold, respectively. A combination of T2 and HT2, T2 and T2-4ol, deoxynivalenol (DON) and nivalenol (NIV), or DON and T2 exhibited synergistic (T2 and HT2 or DON and NIV) or antagonistic (DON and T2) responses.

The ex vivo biophysical response of isolated rat aorta to T-2 toxin, diacetoxyscirpenol (DAS) and Roridin-A were determined. Roridin-A caused a dose-dependent relaxation of precontracted (30 mM KCl) aorta, while T-2 toxin and DAS were without effect. On the other hand, the relaxation response of aorta to isoproterenol and sodium fluoride is impaired in the presence of T-2 toxin or DAS. Roridin-A did not alter the relaxation response to isoproterenol or sodium fluoride. These experiments suggest that macrocyclic trichothecenes (e.g., Roridin-A) are capable of altering vascular smooth muscle tone directly, while simple trichothecenes (e.g., T-2 toxin, DAS) interfere with beta-adrenergic responses.

The toxicity of cultures of Fusarium species and subspecies in section Gibbosum from soils from locations in various climatic regions of Australia was determined using a chick bioassay. Most cultures of F. compactum and F. acuminatum subsp. armeniacum were found to be highly toxic irrespective of geographical origin. The other species and subspecies in section Gibbosum viz; F. acuminatum subsp. acuminatum, F. equiseti, F. scirpi , and F. longipes , were shown to be non-toxic or of low toxicity. Selected culture extracts of F. compactum, F. acuminatum armeniacum and F. acuminatum acuminatum were analysed by gas chromatography after clean up and hydrolysis for the four main trichothecene families, namely; nivalenol (NIV), deoxynivalenol (DON), scirpentriol (Sctol) and T-2 tetraol (T-2tol). Some cultures of F. compactum were found to produce high levels of T-2tol derivatives while others produced high levels of Sctol derivatives. Cultures of F. acuminatum armeniacum produced high levels of T-2tol derivatives and trace levels of Sctol derivatives. In contrast, only trace levels of T-2tol and Sctol were detected in hydrolysed culture extracts of F. acuminatum acuminatum . GC/MS analysis of individual trichothecene derivatives in culture extracts of F. compactum indicated that the main trichothecenes produced were either acuminatin or diacetoxyscirpenol respectively, while F. acuminatum armeniacum was found to produce mainly T-2 toxin and neosolaniol.

Fifty-one species and 3 varieties appertaining to 20 genera were collected from 20 samples of each of hazelnut and walnut seeds on glucose- and 40% (W/V) sucrose-Czapek's agar at 25 degrees C and 45 degrees C with the most common mesophiles were Aspergillus flavus, A. fumigatus, A. niger, Cladosporium cladosporioides, C. herbarum, Penicillium chrysogenum, P. citrinum and P. oxalicum. Fusarium (represented by F. equiseti, F. moniliforme and F. oxysporum) was recovered from walnut seeds in moderate frequency (on glucose-Czapek's agar). Eurotium (E. amstelodami, E. chevalieri, E. repens and E. rubrum) was completely absent on glucose agar, but it was isolated in high frequency from the two types of seeds on 40% sucrose-Czapek's agar. Aspergillus fumigatus and Rhizomucor pusillus were the most common thermophilic fungi in hazelnut and walnut seeds on glucose agar at 45 degrees C. Humicola grisea var. themoidae and Thermoascus aurantiacus were encountered rarely from walnuts. The nuts samples were assayed for natural occurrence of aflatoxins B1, B2, G1 and G2, citrinin, ochratoxin A, patulin, sterigmatocystin, zearalenone, T-2 toxin and diacetoxyscirpenol by thin layer chromatography analysis. Aflatoxin was detected in 90% of hazelnut samples (25-175 micrograms/kg) and 75% of walnut samples (15-25 micrograms/kg). Zearalenone was detected in one sample of walnut (125 micrograms/kg). This is the first report for the presence of zearalenone in walnut. The other mycotoxins were not detected.

Dietary scirpentriol (STO), triacetoxyscirpenol (TAS), monoacetoxyscirpenol (MAS), and diacetoxyscirpenol (DAS), mycotoxins produced by Fusarium species, were compared for their ability to cause mouth lesions when graded dietary levels (0, 1, 2, 4, and 8 micrograms STO or TAS/g; 0, .5, 1, 2, and 4 micrograms MAS or DAS/g) were fed to male broiler chickens for 21 days after hatching. The mouth lesions provoked by each scirpenol were dose-related. The minimum effective doses (MED) were 4, 2, 1, and .5 micrograms/g for TAS, STO, DAS, and MAS, respectively, whether the number of affected birds or the number of affected mouth parts (angles, upper beak, lower beak, and tongue) was the measured response. Lesion sites in the mouth varied with the toxin. The rank orders from greatest to least affected sites were angles, upper beak, lower beak, and tongue for TAS and STO, upper beak, lower beak, angles, and tongue for MAS, and upper beak, lower beak, tongue, and angles for DAS. Mouth lesions were clearly visible with each toxin after feeding for 1 wk and the numbers of affected mouth parts almost tripled after 2 wk exposure. During Week 3 of exposure, only the increase caused by MAS was significant (P less than .05). The MED for growth inhibition were 2, 2, 2, and 8 micrograms/g for STO, MAS, DAS, and TAS, respectively. Thus, mouth lesions were of equal or greater sensitivity than growth inhibition as an indicator of scirpenol toxicity. It would appear that the discovery of mouth lesions in birds justifies a mold and mycotoxin control program.

A quantitative comparison of tissue distribution and excretion of an orally administered sublethal dose of [3H]diacetoxyscirpenol (anguidine) was made in rats and mice 90 min, 24 hr, and 7 days after treatment. Total recoveries of 95–100% were obtained. Approximately 90% of the dose was excreted in urine and feces during the first 24 hr with a feces:urine ratio of about 1:4.5 in both species. Carcass and tissue radioactivity dropped rapidly during the first 24 hr but remained relatively constant at low, but detectable, levels (1.5–3.5% of dose) over the course of the experiment. Few substantive interspecies differences were noted in tissue distribution. At 90 min the highest percentage of dose was in tissues involved in sequestering diacetoxyscirpenol because of high body water/lipid content (carcass, skin) or the absorption (stomach, small intestine), metabolism (liver), or excretion (kidney) of the toxin. The rank order of these tissues was generally stable over the course of the experiment. When data were expressed as specific radioactivity (dpm/g tissue) instead, the carcass and skin dropped from the top rank tissues at 90 min and were replaced by the spleen and cecum. At 24 hr and 7 days the top-ranked order of tissues shifted to include organs associated with trichothecene-induced toxicity such as the lymphohematopoietic system (spleen, thymus, and femur bone marrow), heart, and testis (in mouse) as well as the cecum and large intestine. In addition, the rate of loss of radioactivity with time generally did not decrease as rapidly in these target organs as observed in liver, kidney, skin, and carcass. Brain radioactivity, though very low, also diminished relatively slowly. Significant differences in specific radioactivity which did occur between the rat and mouse tended to occur in target organs and with the higher levels present in the mouse. These data were discussed in terms of interspecies differences in lethality and target organ toxicity.

Scirpentriol (STO) and its seven acetylated derivatives, 3-, 4- and 15-monoacetoxyscirpenol (MAS), 3,4-, 3,15-, and 4,15-diacetoxyscirpenol (DAS), and 3,4,15-triacetoxyscirpenol (TAS) were compared for their acute oral lethality in broiler chicks, lethality in brine shrimp, and dermal toxicity in guinea pigs. Of the eight toxins, 4,15-DAS was the most toxic in the three assays, 3-MAS was the least toxic in brine shrimp and dermal assays, and 3,4-DAS was the least toxic in the chick assay. There was a difference of about a 100-fold and 20-fold, respectively, between 4,15-DAS and 3-MAS in dermal toxicity and brine-shrimp toxicity, as well as a difference of more than 16-fold between 4,15-DAS and 3,4-DAS in chick toxicity. In general, a free hydroxy group at the 3-position was a primary determinant of toxicity. Toxicity in the scirpenol family did not follow precisely the pattern reported earlier for the T-2 toxin family of trichothecene toxins, in which a decrease in the number of acyl groups was accompanied by a decrease in toxicity. At necropsy, the predominant sign in chicks was petechial hemorrhaging, primarily in the gastrointestinal tract and in the vascular beds of the beaks and the toe nails. The 4,15-DAS and 15-MAS were about 3 times more toxic in chicks than aflatoxin. All members of the scirpenol family of trichothecene mycotoxins appeared sufficiently toxic to warrant attention whenever field outbreaks occur. Apparently, brine shrimp and dermal assays are successful predictors of chick lethality by the more toxic trichothecenes and are less suitable for predicting the activity of the less toxic trichothecenes.