No abstract

To test the complex, acute biochemical effects of combined, naturally co‐occurring fusariotoxins, a 5‐day rat study was performed. Mycotoxin treatment was invented by intraperitoneal injection: FB1 (F): 9 µg/animal/day (approx. 30 µg/kg bw/day), DON (D): 16.5 µg/animal/day (approx. 55 µg/kg bw/day) and ZEN (Z): 12.75 µg/animal/day (approx. 42.5 µg/kg bw/day). The binary groups (FB1 and DON [FD], FB1 and ZEN [FZ] and DON and ZEN [DZ]) as well as the ternary (FB1, DON and ZEN [FDZ]) group were dosed at the same combined level as the individual mycotoxins. Body weight, feed intake and mortality were not affected by any of the treatments. FB1 and DON in combination (FD) increased the plasma aspartate aminotransferase activity synergistically (compared to the individual FB1 and DON). In the liver, both the total glutathione (GSH) and the glutathione peroxidase (GPx) activity were increased (p < 0.05) by the binary FB1 and ZEN (FZ) and the DON and ZEN (DZ) groups as well as the ternary FB1, DON and ZEA group (FDZ) compared to the control. The GSH level of the ternary group was significantly increased compared to the FB1 group, whereas the GPx activity of the ternary group was significantly increased compared to all three the individual mycotoxin groups. The Bliss independence method revealed synergism between DON and ZEN (DZ), as well as FB1 and DON (FD) on liver GPx activity. None of the toxins alone or in combination exerted strong genotoxicity on lymphocytes, neither on the gross histopathological characteristics. However, even at these low levels acute exposure of more than one of these mycotoxins (FB1, DON and ZEN) affected metabolic and detoxification changes.

Male Wistar rats were treated intraperitoneally (i.p.) with fumonisin B1 (FB1; 0, 20, 50 and 100 mg/kg dietary dose equivalent) for 5 and 10 days (n = 24–24 in each setting) to gain dose- and time-dependent effects on antioxidant status and oxidative stress response, clinical chemical endpoints and liver, kidney and lung histopathology and lymphocyte damage (genotoxicity). FB1 decreased feed intake, body weight gain and absolute liver weight, irrespective of the toxin dose. Relative kidney weight increased in the 10-day setting. Linear dose response was found for plasma aspartate aminotransferase, alanine aminotransferase, total cholesterol, urea and creatinine, and exposure time-dependence for plasma creatinine level. The latter was coupled with renal histopathological findings, tubular degeneration and necrosis and the detachment of tubular epithelial cells. The pronounced antioxidant response (reduced glutathione accretion, increasing glutathione peroxidase activity) referred to renal cortical response (5–10 days exposure at 50–100 ppm FB1). Hepatic alterations were moderate, referring to initial phase lipid peroxidation (exposure time dependent difference of conjugated diene and triene concentrations), and slight functional disturbance (↑ total cholesterol). Lymphocyte DNA damage was moderate, supporting a mild genotoxic effect of FB1.

Fusarium mycotoxins, such as fumonisin B 1 (FB 1 ), deoxynivalenol (DON) and zearalenone (ZEN), frequently co-occur in feed raw materials and their presence is ubiquitous. The aims of this study were to determine the concentration that inhibits cell viability by 50% (IC 50 values) for each mycotoxin (after 24, 48 and 72 h) and to investigate their combined effects in binary (DON + ZEN: DZ, DON + FB 1 : DF, FB 1 + ZEN: FZ) and ternary (DFZ) mixtures using cyto-and genotoxicity on porcine lymphocytes as endpoints. The potency of cytotoxicity of the three toxins in an increasing order was FB 1 < ZEN < DON. The range of IC values depending on the period of exposure was 0.31-0.42 μg/ml and 16.6-22.9 μg/ml for DON and ZEN, respectively, and 101.15 μg/ml for FB 1 (50% viability was reached only after 72 h). The main interaction observed was antagonism regarding cytotoxicity. Lower and higher sets of concentrations were used for the genotoxicity (comet assay) experiments. When lower concentrations were used, antagonism was again the main interaction observed. However, at higher concentrations an antagonism was confirmed only for DFZ, whereas for DZ and FZ a synergism was observed. Interactions of DF were inconsistent in different exposure periods in both series of experiments. Further studies with additional endpoints should be performed (e.g. DNA fragmentation, protein synthesis) in order to elucidate the mechanisms underlying the interactions observed.

As the most common grain contaminant worldwide, deoxynivalenol is of high importance despite its low toxicity compared to other trichothecene mycotoxins. Data on the effects of deoxynivalenol in rabbits are scarce. Thus, the aim of this study was to investigate the effects of dietary deoxynivalenol fed at a high level (10 mg/kg of feed) on the productive performance, blood indices, immunological variables, histopathological changes, and genotoxicity in rabbits. Forty-eight Pannon White rabbits were exposed to contaminated diets for three weeks. Despite its high concentration, deoxynivalenol did not affect the feed intake, body weight, and body weight gain. Liver and kidney function was not affected, as shown by the clinical chemistry indices. Conversely, in two rabbits the toxin caused mild fibrosis of the liver, without degenerative changes of the hepatocytes. No genotoxicity could be observed either. Gut cytokines and the phagocytic activity of the macrophages did not differ significantly. The percentage of neutrophils was significantly lower, whereas that of eosinophils was significantly higher in the toxin-fed group. Deoxynivalenol did not cause significant changes in gut and villus morphology. In 4 out of the 6 deoxynivalenol-treated animals, the ratio of lymphoblast proliferation and simultaneous apoptosis shifted towards apoptosis in the gut-associated lymphoid tissue. In the central part of the lymphoid follicles of the spleen, lymphocyte depletion and follicular atrophy could be detected. It can be concluded that rabbits are less sensitive to deoxynivalenol, but the findings confirm that this Fusarium toxin is capable of modulating the immune response. Mycotoxin, immune response, histology, blood indicesMycotoxins are secondary metabolites of filamentous fungi occurring worldwide. Despite being the least toxic trichothecene, the ubiquitous occurrence of deoxynivalenol (DON) in grains increases its importance for food and feed safety. The pig is the most sensitive, whereas ruminants are the least sensitive animal species to DON (Pestka and Smolinski 2005;Pestka 2007;Sobrova et al. 2010). Monogastric animals are extremely prone to growth and body weight gain suppression upon DON exposure. Deoxynivalenol possesses immunomodulatory properties as well (Pestka and Smolinski 2005). To the best of our knowledge, only few experiments have been conducted with DON in rabbits (Khera et al. 1986;Hewitt et al. 2012). The aim of this study, therefore, was to investigate the effects of 10 ppm (mg/kg feed) of dietary DON which is twice as much as the highest guidance value

Conventional analytical methods used for the analysis of fumonisin content in animal feeds fail to take into account the fumonisin content bound to the matrix, which is otherwise bioaccessible and can be absorbed from the gastrointestinal tract. Moreover, underestimation of fumonisin content using routine analytical methods can affect animal experiments using cereals contaminated by fungi. In the present study, hidden fumonisin B1 was analysed in two cereal substrates (maize and wheat) which were inoculated with Fusarium verticillioides (MRC 826). The study compared a routine extraction procedure with an in vitro digestion sample pre-treatment. We found that all samples showed a higher content of fumonisin B1 after digestion, compared to the free fumonisin obtained only by extraction. The percentage of the hidden form was 38.6% (±18.5) in maize and 28.3% (±17.8) in wheat, expressed as the proportion of total fumonisin B1. These results indicate that the toxin exposure of the animals determined by the routine fumonisin analysis was underestimated, generally by 40%, as bioaccessibility was not taken into consideration. This is crucial in interpretation (and maybe in re-evaluation) of the results obtained from (other) animal experiments.

Fumonisin-producing fungal species, Fusarium verticillioides, culture was mixed in the diets of 6 piglets for 9 days (Fumonisin B1 [FB1] intake of 17 mg/kg) to investigate whether there is any potential alteration in the caecal bacterial communities between the experimental (with F. verticillioides) and control groups (without F. verticillioides). Plate count agar culturing technique was applied to measure the amount of aerobic and anaerobic bacteria, Escherichia coli, coliforms, Lactobacillus spp. and Clostridium perfringens. A significant difference was observed between the control and experimental group only in the case of aerobic bacteria on Day 4, 8.60 ± 0.22 compared to 8.06 ± 0.20 (P < 0.05), respectively. Quantitative polymerase chain reaction (qPCR) was performed to estimate the DNA copy number of total bacteria, Bacteroides and Prevotella spp., Clostridium spp., E. coli, Enterobacteriales, Firmicutes and Lactobacillus spp. Significant differences were observed between the control and experimental group regarding total bacteria on Day 2 and Day 6, Firmicutes on Day 2 and E. coli and Enterobacteriales on Day 4. Regarding the entire feeding time, no significant difference between the two groups was found in all species of investigated bacteria by the culturing technique and qPCR after an 8-day exposure. The present research contributes to the understanding of how microbiota responds to the FB1 load.

The heat shock protein (Hsp70) level was assessed after 14 days of oral gavage-exposure to fumonisin B1 (FB1: 150 µg/animal/day), deoxynivalenol (DON: 30 µg/animal/day) and zearalenone (ZEN: 150 µg/animal/day), alone or in combinations (in additive manner: FD = FB1 + DON, FZ = FB1 + ZEN, DZ = DON + ZEN and FDZ = FB1 + DON + ZEN) in the liver, kidneys and lung of 24 adult male Wistar rats (n = 3/group). The liver was the most responsive tissue, as compared with kidney and lung. Except of DZ-treatment, mycotoxins elevated the Hsp70 levels in livers. The highest Hsp70-levels (≈ twofold) were in the DON, FD, FZ and FDZ treatments (additive effects). In the kidney, alterations (↑ ≈ twofold) were detected in ZEN, FD, FZ and DZ treatments. The least responsive organ was the lung (↑ only in FDZ, antagonistic effect). DON and ZEA exposures have altered the reduced glutathione concentration (↓) and glutathione peroxidase activity (↓) in the blood serum. The serum malondialdehyde level increased only after exposure to FD (synergistic effect), as compared with the DZ group (antagonistic effect). When the blood clinical chemistry was assessed, significant alterations were in alanine aminotransferase (80% increase in FDZ, antagonistic effect) and total protein (↓ ZEN). Results varied according to the organ, toxin type and interactions. Furthermore, oxidative stress was not the only key player behind the Hsp70 increase, in which another mechanism is suggested.