Determination of deoxynivalenol in cereals by HPLC-UV

A simple method for determination of deoxynivalenol (DON) in cereal samples is described. DON was extracted with methanol, the solvent evaporated, and the residue redissolved with water. This extract was purified on immunoaffinity columns. DON was determined by HPLC with UV-detection. The limits of detection (LOD) and quantification (LOQ) were 10 and 50 μg/kg, respectively. Presented at the 25^th Mykotoxin Workshop in Giessen, Germany, May 19–21, 2003     

A case of sporotrichosis caused by two genetically differentSporothrix schenckii strains

A survey for the natural occurrence of Fusarium mycotoxins, deoxynivalenol (DON), nivalenol (NIV) and zearalenone (ZEN), in Dutch cereals (totaling 29 samples) harvested in 1984/1985, showed that 90%, 79% and 62% of samples were contaminated with DON, NIV and ZEN, respectively. Average contents (ng/g) in the total of positive samples were 221 (DON), 123 (NIV) and 61 (ZEN). Among the cereals examined, the highest concentrations (ng/g) was 3198 (DON), 1875 (NIV) and 677 (ZEN) in a yellow corn sample for animal feed. The results of this survey show that Dutch cereals were relatively significantly contaminated with Fusarium mycotoxins.     

Occurrence of trichothecenes, zearalenone and ochratoxin A in cereals and mixed feed from central Lithuania

A total of 92 samples — 23 winter wheat, 12 summer barley, 5 oats and 52 mixed feed — were collected from a state factory in Kaunas, Lithuania and were analysed for the presence of trichothecenes, zearalenone (ZEN) and ochratoxin A (OA) using gas chromatography with electron capture detection and immunoaffinity column/high performance liquid chromatography with fluorescence and UV detections. Deoxynivalenol (DON), nivalenol (NIV), T-2 toxin and HT-2 toxin were detected at concentrations above 10 μg/kg in 68%, 48%, 38% and 8% of cereal samples, respectively, and in 98%, 88%, 12% and 8% of samples of mixed feed for swine and poultry. More than 10 μg/kg of zearalenone and ochratoxin A were found in 58% and 92% of the mixed feed samples, respectively. The highest concentrations of all analysed trichothecenes in Lithuanian mixed feed and cereal grains, with an exception of T-2 toxin in one oat lot and one sample of mixed feed and OA in two mixed feed samples, were lower than those reported as Lithuanian advisory or tolerance limits.     

Effects of a diet containing fusarium toxins on the fertility of gilts and on bulbourethral gland weight in barrows

Nine gilts weighing 80 kg at the beginning of the trial were fed a mycotoxin contaminated diet containing 2 mg deoxynivalenol (DON) and 0.4 mg zearalenone (ZON) per kg (Diet M). Their daily weight gain until 103 kg BW was reduced in comparison to the nine control animals fed an uncontaminated diet (Diet C) (763 vs. 912 g; p = 0.02). There was no treatment effect on the age at first observed oestrus. Seven and eight gilts receiving Diet M and C, respectively, became pregnant after being mated once or being again mated three weeks later. The examination of the uteri of gilts slaughtered 35–61 days after mating showed that the exposure to DON and ZON had no effect on the number of foetuses per gilt (p = 0.54), but increased their growth rate (p = 0.003). Thus, low dietary DON and ZON levels had no negative effects on the reproductive parameters examined. The hypothesis that the bulbourethral gland weight of barrows can be used for the bioassay of low dietary ZON levels was rejected since feeding Diet M from 80–103 kg BW did not increase the weight of that accessory sex gland (p = 0.51).     

Fusarium toxin residues in physiological samples of piglets

Physiological samples of 100 piglets fed diets containing 0.01, 0.06, 0.15, 0.22 and 0.42 mg ZON and 0.2, 0.8, 1.0, 1.9 and 3.9 mg DON per kg over a period of 35±1.5 days were investigated for concentrations of deoxynivalenol (DON) and zearalenone (ZON) and their metabolites. DON was detected in serum, bile and urine in increasing concentrations corresponding to the diet contamination. The metabolite de-epoxy-DON was detected only in urine. The DON contamination of the diet was closely reflected by the serum concentrations of the piglets. ZON and its metabolite α-zearalenol were detected in bile fluid, liver and urine, while β-zearalenol was only detected in bile fluid. In serum neither ZON nor its metabolites were found. The total concentration of ZON plus its metabolites in the bile fluid corresponded well with the dietary contamination. For all analyses it has to be noted that toxin residues were detectable even in individual samples of piglets fed the control diet containing 0.01 mg ZON/kg and 0.2 mg DON/kg. Presented at the 25^th Mykotoxin Workshop in Giessen, Germany, May 19–21, 2003     

Vorkommen von Deoxynivalenol, Zearalenon und HT-2/T-2 Toxin in Getreide und Getreideerzeugnissen

Since February 2004 in Germany maximum limits forFusarium toxins do exist, while harmonised legislation within the EU was recently published and will come into force in July 2006. Meanwhile, the problematic nature ofFusarium mycotoxins is perceived by all participants of the processing chain of cereals. In this study the presence of deoxynivalenol and zearalenone in comparison to the rarely investigated type A-trichothecenes (HT-2, T-2 toxin) in different cereal-products is discussed. About 1000 cereal-based samples have been analysed using a recently developed multitoxin method based on HPLC-MS/MS technique. Despite, up to now no concrete limit for HT-2/T-2 toxin is discussed, the degree of contamination is of special concern for food products dedicated to be placed on the market, to avoid possible risks for consumers. The used method proved to be extremely sensitive for T-2 toxin with a LOD below 1 μg/kg, therefore a comprehensive data set was achieved.     

A plant virus vector for systemic expression of foreign genes in cereals

Inserts bearing the coding sequences of NPT II and beta-glucuronidase (GUS) were placed between the nuclear inclusion b (NIb) and coat protein (CP) domains of the wheat streak mosaic virus (WSMV) polyprotein ORF. The WSMV NIb-CP junction containing the nuclear inclusion a (NIa) protease cleavage site was duplicated, permitting excision of foreign protein domains from the viral polyprotein. Wheat, barley, oat and maize seedlings supported systemic infection of WSMV bearing NPT II. The NPT II insert was stable for at least 18-30 days post-inoculation and had little effect on WSMV CP accumulation. Histochemical assays indicated the presence of functional GUS protein in systemically infected wheat and barley plants inoculated with WSMV bearing GUS. The GUS constructs had greatly reduced virulence on both oat and maize. RT-PCR indicated that the GUS insert was subject to deletion, particularly when expressed as a GUS-NIb protein fusion. Both reporter genes were expressed in wheat roots at levels comparable to those observed in leaves. These results clearly demonstrate the utility of WSMV as a transient gene expression vector for grass species, including two important grain crops, wheat and maize. The results further indicate that both host species and the nature of inserted sequences affect the stability and expression of foreign genes delivered by engineered virus genomes.     

Two-dimensional environmental profiles of growth,deoxynivalenol and nivalenol production by Fusarium culmorum on a wheat-based substrate

AIMS: To determine the effect of interacting conditions of water activity (aw, 0.99-0.85), temperature (15, 25 degrees C) and time (40 days) on growth and production of the mycotoxins deoxynivalenol (DON) and nivalenol (NIV) by Fusarium culmorum on a wheat-based agar medium. METHODS AND RESULTS: Fusarium culmorum grew optimally at 0.995aw and minimally at 0.90 at both 15 and 25 degrees C. No growth was observed at <0.90aw. Overall, temperature, aw and their interaction had a statistically significant effect on the growth rate of F. culmorum. Production of both DON and NIV were over a much narrower range (0.995-0.95aw) than that for growth. The highest concentrations of DON and NIV levels were produced at 0.995aw and 0.981aw at 25 degrees C, respectively, after 40 days of incubation. Statistically, aw, temperature and incubation time, and aw x temperature and temperature x incubation time had a statistically significant effect on DON/NIV production. CONCLUSIONS: This is the first detailed report on the two-dimensional environmental profiles for DON/NIV production by F. culmorum in the UK. SIGNIFICANCE AND IMPACT OF THE STUDY: As part of a hazard analysis critical control point (HACCP) approach, this type of information is critical in monitoring critical control points for prevention of DON/NIV entering the wheat production chain.     

Influence of organically or conventionally produced wheat on health,performance and mycotoxin residues in tissues and bile of growing pigs

From 1999?–?2001 three different varieties of wheat [Contur (susceptible to Fusarium), Batis and Petrus (less susceptible to Fusarium)] were cultivated under organic and conventional conditions in order to determine mycotoxin burden. Soil quality, preceding crop and weather conditions were comparable in the different production systems. The wheat batches were analysed for moulds, and the contents of zearalenone (ZEN) and deoxynivalenol (DON). Feeding trials were carried out with growing pigs (n?=?96; average initial live weight 22.2 ±?1.5?kg [mean?±?SD]) to examine a possible influence on the animal performance and on mycotoxin residues. The data recorded were clinical conditions, performance, biochemical and hematological data. Residues of ZEN, α- and β-zearalenol (ZEL) and of DON were determined in bile, liver and muscle after slaughtering. Conventionally cultivated wheat was more frequently contaminated with Fusarium and contained more frequently ZEN and DON in higher concentrations than the organically produced wheat. Hematological and biochemical parameters of pigs fed with organically cultivated diets were not different from those of conventionally fed pigs. Pigs fed with organically produced wheat showed a slightly higher daily weight gain, but a lower carcass yield than the conventionally fed animals. The highest residues of DON and total-ZEN (ZEN + α-ZEL + β-ZEL) were found in bile. Bile samples of organically fed pigs contained lower concentrations of total-ZEN than those of conventionally fed pigs. Altogether, these data suggest that wheat from an organic farming does not have higher mycotoxin-contamination than wheat from the conventional farming system.     

Brill-Zinsser disease: report of a case in Canada

This report documents the occurrence of Brill-Zinsser disease in a 48-year-old woman who experienced typhus fever in a German concentration camp. This is the first report of a case of recrudescent typhus in a European immigrating to Canada following World War II. The last reported case of Brill-Zinsser disease in Canada occurred in 1953.     

Effects of a Fusarium toxin-contaminated maize treated with sodium metabisulphite,methylamine and calcium hydroxide in diets for female piglets

Deoxynivalenol (DON) and zearalenone (ZEN) contaminated maize was hydrothermally treated in the presence of sodium metabisulphite (SBS), methylamine and calcium hydroxide (Ca(OH)₂) and included into diets for female piglets to evaluate effects on performance, organ weights, development of hyperestrogenism, serum biochemical parameters, stimulation of peripheral blood mononuclear cells and toxin residues in serum. For this purpose, both uncontaminated maize (CON) and Fusarium toxin-contaminated maize (FUS) were included into diets either untreated (?) or treated (+) according to a 2 by 2-factorial design. One-hundred female weaned piglets were assigned to one of the four treatment groups (n = 25) CON?, CON+, FUS? and FUS+ with DON/ZEN concentrations of 0.43/0.03, 0.04/0.0, 3.67/0.32 and 0.36/0.08 mg per kg diet, respectively. After a feeding period of 27 days, 20 piglets (n = 5) were slaughtered. Performance parameters such as feed intake, live weight gain and feed-to-gain ratio remained unaffected by the treatments. Uterus weights were significantly reduced in group FUS+ compared to FUS? (p = 0.028), while visceral organ weights were not influenced. Vulva width in relation to body weight was highest in group FUS? at the end of the trial, while hydrothermal treatment significantly reduced the parameter (p < 0.01). The highest toxin and toxin metabolite concentrations in serum were detected in group FUS?, whereas ingestion of diet FUS+ reduced the concentrations to the level of the control groups. Serum biochemical and haematological parameters were mainly within the given reference ranges and showed no treatment-related alterations. Stimulation of peripheral blood mononuclear cells was not affected. An effective detoxification of maize by hydrothermal treatment in the presence of SBS, methylamine and Ca(OH)₂ could be demonstrated by means of serum toxin analyses. No undesired side effects of the treated-feed stuff or the chemicals themselves on the health of piglets were detected.     

The effect of iron source on formation and persistence of root iron pools in cereals

Abstract

Five chelated iron sources have been applied to barley and maize to investigate the effect of differing chemical form on the formation and persistence of root apoplastic Fe pools. Short-term Fe exposure (barley) experiments indicated that the charged state of the Fe complex was the most important factor regulating the initial formation and magnitude of the apoplastic pool. Longer term experiments (maize), incorporating a period of Fe deprivation, produced more complex results. Differences in plant growth during the experiment produced changes in the magnitude of the root Fe pool; these interacted with the chemical form of the applied Fe to regulate the release, utilisation and hence the ultimate size of the apoplastic pool produced by each Fe source. It is concluded that such experiments are poor indicators of the potential performance of novel chelated Fe sources.     

A survey of Fusarium toxins in cereal-based foods marketed in an area of southwest Germany

A total of 237 commercially available samples of cereal-based foods including bread and related products, noodles, breakfast cereals, baby and infant foods, rice and other foods were randomly collected in southwest Germany during the first six months of 1998. The trichothecenes deoxynivalenol (DON), 3-and 15-acetyl-deoxynivalenol (3-,15-ADON), nivalenol (NIV), fusarenon-X (FUS-X), T-2 toxin (T-2) and HT-2 toxin (HT-2) were determined by gaschromatography/mass spectrometry following clean-up by a two stage solid-phase extraction. Detection limits ranged between 2 and 12 g/kg. Based on all samples, the incidence of DON, HT-2, T-2, 3-ADON,15-ADON, and NIV was at 71, 18, 4, 4, 4 and 2%, respectively; the average contents in positive samples were at 103, 16, 14, 17, 24 and 109 g/kg,respectively. Fus-X was not detected in any sample. A lower (P < 0.05) DON content was found in baby and infant foods as well as in cookies and cakes compared to bread. Overall, based on the incidence and level of all six toxins, the degree of contamination was lowest in baby and infant foods. Foods produced from either white or whole grain flour did not differ (P > 0.05) with regard to the incidence and level of DON. In foods produced from cereals of organic production both the incidence and median content of DON was lower compared to conventional production. Zearalenone, - and -zearalenol were determined by high performance liquid chromatography in 20 selected samples, mostly baby and infant foods. These toxins were not present in excess of the detection limit in any sample.This revised version was published online in October 2005 with corrections to the Cover Date.     

The influence of rooting density in the soil on P removal by cereals on soils with different yield potential

Summary In field experiments on soils of different rooting density and different yield potential relationships between P uptake and root penetration of the soil, respectively, EUF-P contents in the topsoil were studied.On soils under uniform climatic and management conditions yields increased with increasing total root length in the subsoil (30–100 cm depth), while a negative correlation was found to exist between yields and total root length in the topsoil (0–30 cm).P removal was significantly correlated with the readily available water capacity of the rooted soil volume, which indicates the yield potential of a soil. There was no significant correlation between EUF-P contents of the soils and the P contents of the plants, but P contents in plant dry matter was significantly correlated with rooting density in the topsoil. Considering the effect on P contents of both EUF-P content in the topsoil and rooting density, the correlation coefficient was increased.     

Analysis of theFusarium toxins Deoxynivalenol and Zearalenone in Austrian feeds of the crop years 2002 and 2003

164 feed samples were collected in Lower Austria, Upper Austria, Styria and Carynthia in the crop years 2002 and 2003 and were analysed by means of the RIDASCREEN^® ELISA-assay for theFusarium toxins Deoxynivalenol (DON) and Zearalenone (ZON). The collected samples consisted of swine feeds, cereals and silages. DON concentrations were found up to the range of mg/kg (ppm), in contrast ZON results remained significantly lower.     

Rapid fluorometric test for the quantitative determination of deoxynivalenol in raw cereals

Fast test systems fulfilling legislative specifications are required to determine mycotoxin contamination in unprocessed cerealse.g. at grain elevators, import and export terminals, or the milling and brewing industry to prevent contamination of food and feed. We describe the first tests of a novel fluorescence-based test for deoxynivalenol (DON), which will be commercially available within the next few months. The analytical procedure of the new test takes less than 15 minutes for extraction, purification, derivatization and measurement. The system’s advantage is its speed and easy procedure providing quantitative DON determination. To ensure an international standard, the validation procedure for wheat, barley and maize will be performed following USDA/GIPSA requirements (03/2006) for DON tests. Presented at the 28^th Mykotoxin-Workshop, Bydgoszcz, Poland, May 29–31, 2006 Financial support: Christian Doppler Society and Government of Lower Austria     

Genetic variations in the digestibility in sheep of selected whole-crop cereals used as silages

Whole-plant winter cereals could be of great interest if used as silages for ruminant feeding as opposed to summer crops in that they would spare water resources or valorize low-input management. This study aimed to compare the feeding value of rye, barley, wheat (two genotypes) and triticale (six genotypes). The cereals were sown in October and harvested as silage in June. Forages were offered to Texel castrated sheep in order to evaluate the organic matter digestibility (OMd). The OMd of the wheat cultivars was higher (61.6%, P<0.05) than those of barley (57.2%) and rye (54.7%) but no different from that of triticale (60.6%). Within the triticale genotypes, OMd ranged from 54.7 to 62.3%. The presence of rough barbs should explain the relatively low intake of the cereals with the exception of wheat. Winter cereals provide good-quality forage for feeding ruminants. Wheat has a higher nutritional value than barley and rye and a wide variability for digestibility seems to exist within the triticale cultivars. Such variability in a species known for its ability to be cropped under limiting conditions should be explored in much greater depth as it could result in providing farmers with genotypes of good quality with an acceptable yield at a lower cost.     

Impact of essential oils on growth rate, zearalenone and deoxynivalenol production by Fusarium graminearum under different temperature and water activity conditions in maize grain

AIMS: The effect of five essential oils (oregano, cinnamon, lemongrass, clove and palmarose) on growth rate, zearalenone (ZEA) and deoxynivalenol (DON) production by Fusarium graminearum strains was assessed. METHODS AND RESULTS: The influence of the essential oils was tested on irradiated maize at two concentrations (500 and 1000 mg kg-1), at different water activity (aw) (0.95 and 0.995) and temperature (20 and 30 degrees C) levels. At 0.995 aw all essential oils tested had an inhibitory effect on growth rate of F. graminearum at both temperatures studied. At this aw level, DON production in general was inhibited by all essential oils at 30 degrees C and, although palmarose and clove were the only essential oils with statistically significant inhibitory effect on ZEA production, an inhibitory trend was observed when cinnamon and oregano oils were added to maize grain. CONCLUSIONS: Antifungal and antimycotoxigenic activity of the essential oils assayed was shown to depend on environmental conditions. SIGNIFICANCE AND IMPACT OF THE STUDY: It is apparent that essential oils should be considered as alternative preharvest natural fungicides. Further investigation on natural maize grain might be useful to study the effectiveness of these essential oils in the presence of natural mycoflora of maize grain.     

HPLC-PAD-APCI/MS assay of phenylpropanoids in cereals

A new, rapid HPLC-PAD-APCI/MS assay has been developed in order to measure accurately the amount of p-coumaric, E- and Z-ferulic acid and the dehydrodimers of ferulic acid in cereal grain. In the positive ionisation mode, MS patterns gave additional information for the identification of the dimers. The time required and the quantities of solvents employed in the developed analytical method are much lower than those involved in previously available assays of these compounds, thus making the method suitable for the screening of cereal genotypes. Application of the method to accessions of maize, wheat and sorghum showed that E-ferulic was the most abundant phenylpropanoid, whilst the major dimer was 8-O-4' dehydrodimer of ferulic acid followed by the 5-5' and then the 8-5' forms. Maize grains, especially of the Mexican landraces, contained the highest levels of these dimers.