The effects of the Penicillium mycotoxins citrinin, cyclopiazonic acid, ochratoxin A, patulin, penicillic acid, and roquefortine C on in vitro proliferation of porcine lymphocytes

The in vitro effect of each of the Penicillium mycotoxins citrinin (CIT), cyclopiazonic acid (CPA), ochratoxin A (OTA), patulin (PAT), penicillic acid (PIA) and roquefortine C (RQC) on mitogen induced lymphocyte proliferation was determined using purified lymphocytes from 6 piglets. Dose response curves for each mycotoxin were generated and the concentrations producing 50% inhibition of cell proliferation (IC₅₀) were estimated. OTA and PAT were the most potent toxins with IC₅₀ of 1.3 and 1.2 μmol/l, respectively (0.52 and 0.18 mg/l, respectively). Based on molar concentrations, OTA was 15, 30, 40, and 65 times more potent as an inhibitor than PIA, CIT, CPA and RQC, respectively.     

Natural multi-occurrence of mycotoxins in rice from Niger State,Nigeria

Twenty-one rice samples from field (ten), store (six) and market (five) from the traditional rice-growing areas of Niger State, Nigeria were analysed for aflatoxins (AFs), ochratoxin A (OTA), zearalenone (ZEA), deoxynivalenol (DON), fumonisin B₁ (FB₁) and B₂ (FB₂), and patulin (PAT) by thin-layer chromatography (TLC) and high-performance liquid chromatography (HPLC) respectively. T-2 toxin was determined using TLC only. AFs were detected in all samples, at total AF concentrations of 28–372 μg/kg. OTA was found in 66.7% of the samples, also at high concentrations (134–341 μg/kg) that have to be considered as critical levels in aspects of nephrotoxicity. ZEA (53.4%), DON (23.8), FB₁ (14.3%) and FB₂ (4.8%) were also found in rice, although at relatively low levels. T-2 toxin was qualitatively detected by TLC in only one sample. Co-contamination with AFs, OTA, and ZEA was very common, and up to five mycotoxins were detected in a single sample. The high AF and OTA levels as found in rice in this study are regarded as unsafe, and multi-occurrences of mycotoxins in the rice samples with possible additive or synergistic toxic effects in consumers raise concern with respect to public health.     

Mycotoxins in horse feed

A total of 62 samples of commercial horse feed preparations (complementary feeds) containing cereal mixtures (“muesli” or mash, n = 39; pelleted feeds, n = 12), and plain horse feed grains (maize, n = 5; oats, n = 4; barley, n = 2) were purchased from 21 different producers/distributors from the German market. All samples were analysed by competitive enzyme immunoassays (EIA) for six different mycotoxins (mycotoxin groups). Analytes (detection limit, mean recovery) were: deoxynivalenol (DON, 10 μg/kg, 84%), zearalenone (ZEA, 5 μg/kg, 93%), fumonisin B₁ (FB₁, 2 μg/kg, 113%), T-2 toxin (T-2, 0.1 μg/kg, 71%), sum of T-2 + HT-2 toxin (T-2/HT2, 0.2 μg/kg, 97%), ochratoxin A (OTA, 0.2 μg/kg, 67%), and total ergot alkaloids (Generic Ergot Alkaloids “GEA”, 30 μg/kg, 132%). All samples contained DON (16–4,900 μg/kg, median 220 μg/kg), T-2/HT-2 (0.8–230 μg/kg, median 24 μg/kg), and T-2 (0.3–91 μg/kg, median 7 μg/kg). ZEA was detected in 98% of the samples (7–310 μg/kg, median 61 μg/kg). Most samples (94%) were positive for FB₁ (2–2,200 μg/kg, median 27 μg/kg). Ergot alkaloids were detected in 61% of samples (28–1,200 μg/kg, median 97 μg/kg), OTA was found in 42% of samples (0.2–4 μg/kg, median 0.35 μg/kg). The results demonstrate that a co-contamination with several mycotoxins is very common in commercial horse feed from the German market. The toxin concentrations were in most cases well below the levels which are usually considered as critical or even toxic. The highest mycotoxin concentrations were mostly found in single-grain cereal feed: the maximum values for DON and FB₁ were found in maize, the highest T-2/HT-2 toxin concentrations were found in oats, and the highest concentration of ergot alkaloids was found in barley. In composed feeds, no correlation between cereal composition and mycotoxin levels could be found.     

Exposure to Penicillium mycotoxins alters gene expression of enzymes involved in the epigenetic regulation of bovine macrophages (BoMacs)

In this study, the modulation of key enzymes involved in epigenetic regulation was assessed in immortalized bovine macrophages (BoMacs) following in vitro exposure to the following Penicillium mycotoxins: citrinin (CIT), ochratoxin A (OTA), patulin (PAT), mycophenolic acid (MPA), penicillic acid (PA), or a combination of one of the above with OTA at the concentration that inhibits BoMac proliferation by 25 % (IC25). Real-time PCR analysis of the genes coding DNA methyltransferases (DNMTs), histone demethylases (JMJD-3 and UTX), as well as the class-1 histone deacetylases (HDAC?1, ?2, and ?3) and histone acetylase (Bmi-1) was assessed following 6 h of mycotoxin exposure. A change in the expression of JMJD-3 as well as HDAC-3, MPA (p?=?0.1) and PA (p?=?0.08), by at least one of the treatments was observed at their respective IC25. The expression of JMJD-3 was significantly induced by PA, but synergistically suppressed by CIT?+?OTA. The combination of CIT?+?OTA also synergistically suppressed the expression of DNMT-3a and DNMT-3b. The combination of PAT?+?OTA reduced DNMT-3a expression, while PA?+?OTA reduced DNMT-3b expression. Lastly, MPA and PA slightly reduced HDAC-3 expression, while OTA in combination with CIT, PAT, MPA or PA synergistically suppressed HDAC-3 expression. The results of this study demonstrate that Penicillium mycotoxin exposure, specifically OTA and other mycotoxin combinations, can alter the expression of BoMac enzymes that are involved in epigenetic regulation. These findings suggest a potential novel regulatory mechanism by which mycotoxins can modulate macrophage function.     

Combinatory effects of citrinin and ochratoxin A in immortalized human proximal tubule cells

Ochratoxin A (OTA) and citrinin (CIT) are two mycotoxins often occurring together in grains and cereals. Although both are nephrotoxic and can induce apoptosis, combination effects have not been examined up to now. Therefore, the aim of this study was to take a close look at the interactions of citrinin and OTA in cultured human proximal tubule-derived cells (IHKE cells). The cytotoxicity of both mycotoxins was studied, measuring the metabolic activity and the cell number. Furthermore, caspase 3-activation as a marker for apoptosis was examined for both mycotoxin alone and in combination. The results show that citrinin had an antagonistic effect on ochratoxin A induced caspase 3-activation in concentrations of 2.5 and 5 μmol/l. Higher concentrations (7.5 and 15 μmol/l) lead to additive effects, lower citrinin concentrations (0.25 and 1 μmol/l) did not show any effect at all. The observed decrease in caspase 3-activity was specific for the combination with OTA, since the combination of citrinin with cisplatin did not show any effect. Citrinin did not influence of the OTA-induced apoptosis when added two hours after applying ochratoxin A. Also the combination of both toxins decreased the uptake of OTA into the cells which might be an explanation for the antagonistic effect of citrinin in certain concentrations. However, the transport into cells can not be the only explanation. so further examinations are necessary. Presented at the 27^th Mykotoxin-Workshop. Dortmund, Germany, June 13–15, 2005.     

Impact of some environmental factors on growth and production of ochratoxin A of/by <Emphasis Type="Italic">Aspergillus tubingensis,A. niger</Emphasis>, and <Emphasis Type="Italic">A. carbonarius</Emphasis> isolated from moroccan grapes

The effects of temperature, water activity (a_w), incubation time, and their combinations on radial growth and ochratoxin A (OTA) production of/by eight Aspergillus niger aggregate strains (six A. tubingensis and two A. niger) and four A. carbonarius isolated from Moroccan grapes were studied. Optimal conditions for the growth of most studied strains were shown to be at 25°C and 0.95 a_w. No growth was observed at 10°C regardless of the water activity and isolates. The optimal temperature for OTA production was in the range of 25°C∼30°C for A. carbonarius and 30°C∼37°C for A. niger aggregate. The optimal a_w for toxin production was 0.95∼0.99 for A. carbonarius and 0.90∼0.95 for A. niger aggregate. Mean OTA concentration produced by all the isolates of A. niger aggregate tested at all sampling times shows that maximum amount of OTA (0.24 μg/g) was produced at 37°C and 0.90 a_w. However, for A. carbonarius, mean maximum amounts of OTA (0.22 μg/g) were observed at 25°C and 0.99 a_w. Analysis of variance showed that the effects of all single factors (a_w, isolate, temperature and incubation time) and their interactions on growth and OTA production were highly significant.     

Guttation droplets of <Emphasis Type="Italic">Penicillium nordicum</Emphasis> and <Emphasis Type="Italic">Penicillium verrucosum</Emphasis> contain high concentrations of the mycotoxins ochratoxin A and B

Eight of eleven ochratoxigenic isolates of Penicillium nordicum and Penicillium verrucosum produced guttation droplets when grown on Czapek yeast extract (CYA) agar for 10–14 days at 25°C. Parallel cultivation of one strain each of P. nordicum and P. verrucosum on malt extract agar demonstrated that higher volumes of exudate are produced on this agar. However, HPLC analyses revealed higher concentrations of ochratoxin A (OTA) and B (OTB) in droplets originating from cultures on CYA. For quantitative determination of the mycotoxin contents, triplicates of three isolates each of P. nordicum and P. verrucosum were grown as single spot cultures on CYA for up to 14 days at 25°C. Guttation droplets were carefully collected between day 11 and 14 with a microliter syringe from each culture. Extracts from exudates and corresponding mycelia as well as fungal free agar were analyzed by HPLC for the occurrence of ochratoxin A (OTA) and B (OTB). Mean concentrations ranging between 92.7–8667.0 ng OTA and 159.7–2943.3 ng OTB per ml were detected in the guttation fluids. Considerably lower toxin levels were found in corresponding samples of the underlying mycelia (9.0–819.3 ng OTA and 4.5–409.7 ng OTB/g) and fungal free agar (15.3–417.0 ng OTA and 12.7–151.3 ng OTB/g). This is the first report which shows that high amounts of mycotoxins could be excreted from toxigenic Penicillium isolates into guttation droplets.     

Toxicity of ochratoxin A in aBrevibacillus brevis — Growth inhibition assay

Ochratoxin A (OTA) is a nephrotoxic, carcinogenic and immunosuppressive mycotoxin. It can be detoxified by various microorganisms, e.g. different yeast strains, via metabolisation into ochratoxin α (OTα). Within this study a growth inhibition assay was developed to compare the toxicity of OTA and its degradation product OTα. As an indicator organismBrevibacillus brevis was used. The assay was performed in microtiterplates. Growth inhibition was determined by comparing the optical density values ofBrevibacillus brevis cultures grown in medium supplemented with OTA/OTα and OTA/OTα-free medium, respectively. It could be shown thatB. brevis is sensitive to OTA (EC₁₀₀=0.5 mg/L±0.03 mg/L), which is not the case for its metabolite OTα. Therefore this bioassay is a useful tool to show the detoxification of OTA to OTα by microbial degradation. Presented at the 25^th Mykotoxin Workshop in Giessen, Germany, May 19–21, 2003     

Mycotoxin Ochratoxin A-induced cell death and changes in oxidative metabolism of Arabidopsis thaliana

We evaluated the phytotoxicity of mycotoxin ochratoxin A (OTA) from Aspergillus and Penicillium strains on Arabidopsis thaliana. The results demonstrate that the growth of Arabidopsis thaliana on media containing OTA was inhibited significantly. Moreover, OTA induced necrotic lesions in detached leaves, which are reminiscent of hypersensitive response lesions that are activated during plant–pathogen interactions and other abiotic stress factors. From our study, we can see that OTA exposure stimulated a biphasic oxidative burst in the leaves, resulting in the generation of hydrogen peroxide (H₂O₂) and superoxide anion radicals (O₂^·−) and in the concomitant down-regulation of antioxidant enzyme defense responses and up-regulation of lipid peroxidation. These results suggested that OTA damage might result from reactive oxygen species pathways. Our experiments provide a useful model plant system for research on OTA-induced plant cell death.     

Adsorption of ochratoxin A and zearalenone by potential probiotic Saccharomyces cerevisiae strains and its relation with cell wall thickness

Aims: To examine Saccharomyces cerevisae strains with previously reported beneficial properties and aflatoxin B₁ binding capacity, for their ability to remove ochratoxin A (OTA) and zearalenone (ZEA) and to study the relation between cell wall thickness and detoxificant ability of yeast strains. Methods and Results: A mycotoxin binding assay at different toxin concentrations and the effect of gastrointestinal conditions on mycotoxin binding were evaluated. Ultrastructural studies of yeast cells were carried out with transmission electronic microscopy. All tested strains were capable of removing OTA and ZEA. Saccharomyces cerevisiae RC012 and RC016 showed the highest OTA removal percentage, whereas RC009 and RC012 strains showed the highest ZEA removal percentages. The cell diameter/cell wall thickness relation showed a correlation between cell wall amount and mycotoxin removal ability. After exposure to gastrointestinal conditions, a significant increase in mycotoxin binding was observed. Conclusions: All tested Saccharomyces cerevisiae strains were able to remove OTA and ZEA, and physical adsorption would be the main mechanism involved in ochratoxin A and ZEA removal. Gastrointestinal conditions would enhance adsorption and not decrease mycotoxin–adsorbent interactions. Significance and Impact of the Study: Live strains with mycotoxin binding ability and beneficial properties are potential probiotics that could be included in animal feed. Previous and present results suggest that the RC008 and RC016 strains are very promising candidates for functional feed product development.     

No OTA in fresh cocoa beans

The presence of ochratoxin A (OTA) in cocoa and chocolates has been reported. There is no previously published data available on the source and development of OTA producing moulds and OTA itself in cocoa,i.e. where the mycotoxin enters the cocoa supply chain. A selection of fresh and undamaged cocoa pods from various growing regions was examined for mycotoxin OTA content. In addition, a small selection of damaged or mouldy cocoa pods was included in the examination. It was shown that the ripening phase of healthy cocoa pods from the tree up to being harvested was not a critical period for the occurrence of the mycotoxin OTA. The mycotoxin OTA was not detectable in any of the analysed cocoa pods. Presented at the 28^th Mykotoxin-Workshop, Bydgoszcz, Poland, May 29–31, 2006     

Programmed cell death by interaction of ochratoxin A with other nephrotoxins

We determined the effects of co-exposure of OTA and other (potentially) nephrotoxic substances on renal tubule cells (IHKE, from the proximal tubule; MDCK-C7 cells, representing principal cells of the collecting duct) by measuring the activation of caspase-3, an enzyme with key roles in the process of programmed cell death or apoptosis.Substances used in combination with OTA included: cadmium, a known nephrotoxin with apoptotic effects, cisplatin (anticancer drug, also a known nephrotoxin), cyclosporin A (an immunosuppresive agent), H₂O₂ (generated during oxidative cell stress), amphotericin B (antifungal agent), and fumonisin B1. Because of the great number of possible combinations, a method based on the 96-well format was developed which allowed a fast and cost-efficient succesive measurement of caspase-3 activity and protein content in one well as well as necrotic effects in parallel.We found that cells responded differently to the various combinations: for example, 1 µM OTA combined with 100 µM cisplatin had an antagonizing effect on caspase-3 activation in IHKE cells but potentiating effects in MDCK-C7. DNA ladder formation confirmed the results. Necrotic effects were very small and additive. We conclude that the co-exposure of renal cells to OTA with other substances can enhance or reduce the apoptotic potential of one substance alone depending on the substance and on the cell line investigated. A “harmless” substance can thus convert to a potent cell toxic substance when combined with OTA or other mycotoxins.These first findings show the necessity to further investigate the combined effects of OTA or other mycotoxins with other substances or of mycotoxins with each other.     

Repair of ochratoxin A-induced DNA damage and modulation of OTA-related genotoxicity by co-incubation with bile acids and methotrexatein vitro

The mycotoxin ochratoxin A (OTA) induced DNA strand breaks in porcine urinary bladder epithelial cells (PUBEC) and in Madin Darby canine kidney (MDCK) cells. A co-incubation with bile acids or methotrexate reduced or even prevented this adverse effect of OTAin vitro. The protective effect is possibly attributable to a decreased OTA uptake in cells, since bile acids and methotrexate are known to share common transport systems such as organic anion transporters (OAT) and/or organic anion transporting polypeptides (OATP) with the mycotoxin. OTA uptake in cells and its modulation can be one factor which determines the extent of adverse effects in different cell types. Another aspect of interest in this regard relates to repair of DNA damage: PUBEC cells are sensitive to OTA-induced damage which is more pronounced when DNA repair is blocked (by cytosine β-D-arabino-furanosid/hydroxyurea). On the other hand, when cells are kept in fresh (toxin-free) medium for 3 h, OTA-induced DNA damage decreased to control levels. Presented at the 26^th Mykotoxin-Workshop in Herrsching, Germany, May 17–19, 2004     

Mycotoxigenic fungi in peanuts from different geographic regions of Egypt

To understand the importance of mycotoxigenic fungi in Egyptian peanuts, samples from five regions (Alexandria, El-Beheira, El-Sharqiya, El-Daqahelaya in northern Egypt and Asyut, southern Egypt) in two seasons (2007, 2008) were collected. Aspergillus was consistently the most frequent genus in seeds and in-shell peanuts and was the dominant mycotoxigenic component of the mycobiota. There was no direct correlation between the moisture content of the samples and the fungal populations on peanut seeds tested from different regions. The most common species were from Aspergillus section Flavi (4.7-78.3%), Aspergillus section Nigri (9.4–52.6%) and Aspergillus section Circumdati (5.1–30.9%). In the in-shell peanut samples, the lowest populations were recorded in El-Beheira and Asyut (3.7–4.0 log₁₀ CFU g^-1) and the highest in Alexandria and Elsharqiya (4.1–6.0 log₁₀ CFU g^-1). Aspergillus section Flavi and section Nigri were the most dominant, and Aspergillus section Circumdati were only found in samples in 2008. Both qualitative (coconut cream agar) and quantitative analyses (HPLC) were used to analyse the potential mycotoxin production by strains isolated from peanuts. Of a total of 88 Aspergillus section Flavi strains examined, 95% were A. flavus based on production of aflatoxin B₁ on yeast extract sucrose (YES) medium and confirmation using molecular analyses. Of 64 Aspergillus section Circumdati strains only 28% produced ochratoxin A (OTA), and were identified as A. westerdijkiae. No Aspergillus section Nigri strains produced OTA, and they were identified as A. niger (uniseriate). The presence of these toxigenic fungi indicates that there is a potential risk of mycotoxin contamination in Egyptian peanuts and suggests that problems can arise from contamination with both aflatoxins and perhaps also OTA.     

Nitrogen and fungicide applications against Erysiphe cruciferarum affect quality components of oilseed rape

Oilseed rape (Brassica napus L.) is one of the most important oilseed crops in temperate climates. Erysiphe cruciferarum is an important disease of oilseed rape and causes crop loss in warmer areas of Europe. The research investigated the effect of nitrogen fertilizer and fungicidal treatment against powdery mildew infection caused by E. cruciferarum of oilseed rape on seed components, including protein, oil, oleic acid, linolenic acid and undesirable substances such as sinapic acid esters (SAE) and glucosinolates (GSL), using near infrared spectroscopy (NIRS). Five susceptible oilseed rape varieties were employed in this research using four treatment groups: no nitrogen fertilization and no fungicidal treatment (N₀–F₀); no nitrogen fertilization but fungicidal treatment (N₀–F₁); and nitrogen fertilization but no fungicidal treatment (N₁–F₀); nitrogen fertilization and fungicidal treatment (N₁–F₁). Nitrogen fertilization increased the protein, but lowered the oil content, of the seeds. Fungicidal treatments significantly increased oil contents in all varieties tested, however reduced protein levels in fertilized and non-fertilized plots. The level of linolenic acid did not change significantly in any plots of any treatment combinations; a similar result was observed in the level of oleic acid in most of the genotypes. Nitrogen fertilization increased GSL and SAE levels, whereas fungicidal treatment had no effect. Our findings demonstrated that nitrogen fertilization can markedly influence some quality parameters in oilseed rape; however, the application of fungicides reduced side effects of nitrogen fertilizer and resulted a reduction on GSL, SAE and protein contents but an increase on total oil and oleic acid contents.     

Allium cepa as a biomonitor of ochratoxin A toxicity and genotoxicity

Ochratoxin A (OTA) is a toxin produced by Aspergillus and Penicillum moulds. Since OTA has not yet been evaluated in plant systems, this paper focused on describing the controversial effect OTA in an Allium root test model, which has known sensitivity to genotoxins and could be useful in toxin screening. Analyses of root growth and the root meristematic zone in response to OTA treatment were undertaken. The results show OTA toxicity to root growth at a concentration of 10 ug·ml^?1 associated with inhibition of proliferation activity. Cytological changes observed in the Allium chromosome aberrations assay, at a concentration of 5.0 ug·ml^?1, showed that OTA was able to induce genotoxicity at the chromosome level. These results indicate that plants cells (Allium cepa) are very sensitive to the mycotoxin OTA, as observed at the highest concentration. Under these conditions, OTA produced toxicity and cytogenetic injury. Evidence in vitro and in vivo indicates that OTA can induce damage at the DNA level.     

Natural Occurrence of Mycotoxins in Staple Cereals from Ethiopia

The occurrence of mycotoxins in barley, sorghum, teff (Eragrostis tef) and wheat from Ethiopia has been studied. Samples were analyzed for aflatoxin B₁ (AFB1), ochratoxin A (OTA), deoxynivalenol (DON), nivalenol (NIV) and zearalenone (ZEN) using high performance liquid chromatography (HPLC) and for fumonisins (FUM) using enzyme linked immunosorbent assay (ELISA). AFB1 and OTA were detected in samples of all the four crops. AFB1 was detected in 8.8% of the 352 samples analyzed at concentrations ranging from trace to 26 μg kg⁻¹. OTA occurred in 24.3% of 321 samples at a mean concentration of 54.1 μg kg⁻¹ and a maximum of 2106 μg kg⁻¹. DON occurred in barley, sorghum and wheat at 40–2340 μg kg⁻¹ with an overall incidence of 48.8% among the 84 mainly ‘suspect’ samples analyzed; NIV was co-analyzed with DON and was detected at 40 μg kg⁻¹ in a wheat sample and at 50, 380, and 490 μg kg⁻¹ in three sorghum samples. FUM and ZEN occurred only in sorghum samples with low frequencies at concentrations reaching 2117 and 32 μg kg⁻¹, respectively. The analytical results indicate higher mycotoxin contamination in sorghum, which could be related to the widespread storage of sorghum grain in underground pits leading to elevated seed moisture contents. This is the first report on the occurrence of OTA in teff.     

The nephrotoxin ochratoxin A induces apoptosis in cultured human proximal tubule cells

To test the apoptotic potential of the nephrotoxic mycotoxin ochratoxin A (OTA), we exposed human proximal tubule-derived cells (IHKE cells) for various times to OTA concentrations close to those occurring during dietary exposure (from 2 to 100 nmol/L) and investigated caspase 3 activation, chromatin condensation, and DNA fragmentation. OTA induced a time- and concentration-dependent activation of caspase 3: concentrations as low as 5 nmol/L OTA caused a slight but significant increase in caspase 3 activity after 7 days of OTA exposure. Exposure to 10 nmol/L OTA for 72 or 24 h led to a significantly increased activity of caspase 3 in human proximal tubule-derived cells. Radical scavengers such as N-acetylcysteine had no effect on OTA-induced caspase 3 activation. Chelation of intracellular calcium with 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid tetrakis (acetoxymethylester) (BAPTA-AM) also showed no effect. Exposure to 30 nmol/L or more OTA led to DNA fragmentation and chromatin condensation in IHKE cells. Cultured renal epithelial MDCK-C7 and MDCK-C11 or OK cells also showed increased caspase 3 activity after OTA exposure. We conclude that exposure to low OTA concentrations can lead to direct or indirect caspase 3 activation and subsequently to apoptosis in cultured human proximal tubule cells and in other renal epithelial cell lines of different origins. This revised version was published online in July 2006 with corrections to the Cover Date.     

Effect of ochratoxin A and ochratoxin C on the monocyte and lymphocyte function

The effect of practically relevant mycotoxin concentrations on functions of immune cells was studied in in vitro experiments. Porcine mononuclear cells were exposed to a crudeAspergillus-ochraceus toxin containing OTA, a HPLC fraction identical with OTC derived from the crude toxin (RE2), as well as pure OTA and OTC in a concentration range from 0.46 to 3000 ng/ml. The influence of mycotoxin exposure on metabolic activity, mitogen induced proliferation, expression of the activation marker CD25 and the cell cycle of lymphocytes and on the formation of free oxygen radicals as well as the production of the cytokines IL-6 and TNF-α by monocytes was determined. Exposure to high concentrations of all mycotoxin preparations lead to non-specific suppression of the immune cell functions, which was related to cytotoxic effects. Low concentrations caused ambivalent reactions, especially on monocyte function. In general, the HPLC fraction RE2 had an up to 100-fold stronger effect than pure OTA. Ochratoxin-induced suppression of lymphocyte proliferation was not abrogated by phenylalanine or aspartame. The results indicate that immunomodulation can be caused by very low mycotoxin concentrations which are not related to clinical symptoms or loss of performance.