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Little is known about the effects of Fusarium mycotoxins on the fermentation potential of rumen fluid sampled from lactating dairy cows ingesting diets contaminated at regular levels of these mycotoxins (i.e., contamination levels that can normally be found on dairy farms). In the current experiment, rumen donor animals received diets contaminated with both deoxynivalenol (DON) and fumonisins (FB) with or without a mycotoxin-deactivating product. The rumen fluid donor animals were 12 lactating Holstein dairy cows that received one of 3 experimental diets in agreement with a 3 × 3 Latin square design (3 periods and 3 treatments). The 3 diets were as follows: (1) a TMR contaminated with a regular level of Fusarium mycotoxins [340.5 ± 161.0 µg of DON/kg of dry matter (DM) and 127.9 ± 43.9 µg of FB/kg of DM; control diet, CTR], (2) a TMR contaminated with Fusarium mycotoxins at levels higher than CTR but below US and European Union guidelines (733.0 ± 213.6 µg of DON/kg of DM and 994.4 ± 323.2 µg of FB/kg of DM; MTX), and (3) the MTX diet (897.3 ± 230.4 µg of DON/kg of DM and 1,247.1 ± 370.2 µg of FB/kg of DM) supplemented with a mycotoxin-deactivator product (Mycofix, Biomin Holding GmbH; 35 g/animal per day; MDP). Each experimental period lasted 21 d, and rumen fluid was individually sampled from all cows on the last day of each intoxication period. Then, the 4 rumen fluids sampled from cows receiving the same experimental diets were pooled into a single rumen inoculum, which was used in the in vitro gas production test. For the gas production test, 3 different rumen inocula (i.e., CTR, MTX, and MDP) were buffered (buffer:rumen ratio of 2:1, vol/vol) and then used in 3 fermentation runs to evaluate gas production dynamics in the presence of 8 feeds (i.e., corn meal, barley meal, corn silage, sorghum silage, alfalfa hay, ryegrass hay, dry brewers barley grains, and dried distillers grains with solubles). The kinetic parameters of gas production and volatile fatty acid concentrations were evaluated at the end of fermentation. The block run (i.e., fermentation day) effect influenced all of the fermentative and kinetic parameters. Greater final volumes or rates of gas production over time were observed for MDP compared with MTX rumen inocula (i.e., 172.6 vs. 147.8 mL/g of organic matter or 0.078 vs. 0.063 h−1, respectively). However, the increase in rate of gas production was not consistent among tested feeds, meaning that a treatment by feed interaction was observed. Volatile fatty acid concentrations were not different among treatments, except for a slight increase of acetic acid in CTR compared with MTX (i.e., 71.0 vs. 67.9 mmol/L). This study showed that Fusarium-produced mycotoxins negatively affected the kinetics of gas production in feeds, whereas the presence of the mycotoxin-deactivator product in the diets of donor animals resulted in an increase in rumen fermentation potential, thus safeguarding the rumen environment.

Interactions between livestock management practices and toxicological outcomes of mycotoxin exposure may explain the range of tolerable toxin levels reported for various species. In the current study, we investigated the effect of concurrent mycotoxin exposure with a high starch diet in 12 beef steers in a partial cross-over experiment using a 21-day treatment period, followed by a 14-day clearance. During the treatment period, animals were assigned to one of two diets: a low mycotoxin control total mixed ration (TMR) (0.2±0.1 mg deoxynivalenol (DON) and 0.2±0.2 mg fumonisins (FUM)/kg TMR) and a high mycotoxin TMR treatment (1.7±0.2 mg DON and 3.5±0.3 mg FUM/kg TMR). We evaluated the impacts of these mycotoxins on performance, physiology and biochemistry; and the ability of the clearance period to return animals to a naïve state in the cross-over model. The lack of acute ruminal acidosis observed indicates that the animals were able to withstand the physiological stresses of the high starch diet, while toxicological outcomes were manifested in minor perturbations of biochemistry and outright performance of exposed animals. Aspartate aminotransferase, cholesterol, fibrinogen and leukocyte count were increased while sorbitol dehydrogenase, bile acids and mean corpuscular volume were decreased in treatment-fed steers, yet were not significantly different than those from control-fed animals. Fusarium toxin exposure significantly decreased ruminal fluid pH, with the clearance period returning animals to a naïve state, as it did for most of the molecular variables measured. Conversely, treatment-fed animals continued to exhibit significantly lower average weekly body weight throughout the treatment period and the first week of the clearance period. While the risk of adverse health effects to fattening cattle from similar doses of DON or FUM as used in the current study is considered low, additional work should be directed towards minimising production losses due to these feed contaminants.

A common problem that arises from the use of plant ingredients is the presence of mycotoxins. The plant meals with more potential to be used in turbot diets have a natural tendency to be contaminat...

The main objective of this study was to evaluate the effects of sodium sulfite (SoS) treatment of maize and its impact on the porcine immune system in the presence of an LPS-induced systemic inflammation. Control maize (CON) and Fusarium-toxin contaminated maize (FUS) were wet-preserved (20% moisture) for 79 days with (+) or without (−) SoS and then included at 10% in a diet, resulting in four experimental groups: CON−, CON+, FUS−, and FUS+ with deoxynivalenol (DON) concentrations of 0.09, 0.05, 5.36, and 0.83 mg DON/kg feed, respectively. After 42-day feeding trial (weaned barrows, n = 20/group), ten pigs per group were challenged intraperitoneally with either 7.5 μg LPS/kg BW or placebo (0.9% NaCl), observed for 2 h, and then sacrificed. Blood, mesenteric lymph nodes, and spleen were collected for phenotyping of different T cell subsets, B cells, and monocytes. Phagocytic activity and intracellular formation of reactive oxygen species (ROS) were analyzed in both polymorphonuclear cells (PMN) and peripheral blood mononuclear cells (PBMC) using flow cytometry. Our results revealed that the impact of DON was more notable on CD3+CD4+CD8+ T cells in lymphoid tissues rather than in blood T cells. In contrast, SoS treatment of maize altered leukocyte subpopulations in blood, e.g., reduced the percentage and fluorescence signal of CD8high T cells. Interestingly, SoS treatment reduced the amount of free radicals in basal ROS-producing PMNs only in LPS-challenged animals, suggesting a decrease in basal cellular ROS production (pSoS*LPS = 0.022).

International audience; The impact of the Fusarium-derived metabolites beauvericin, enniatin B and B1 (EB) alone or in combination with deoxynivalenol (DON) was investigated in 28-29 days old weaning piglets over a time period of 14 days. The coapplication of EB and DON (EB + DON) led to a significant decrease in the weight gain of the animals. Liver enzyme activities in plasma were significantly decreased at day 14 in piglets receiving the EB + DON-containing diet compared to piglets receiving the control diet. All mycotoxin-contaminated diets led to moderate to severe histological lesions in the jejunum, the liver and lymph nodes. Shotgun metagenomics revealed a significant effect of EB-application on the gut microbiota. Our results provide novel insights into the harmful impact of emerging mycotoxins alone or with DON on the performance, gut health and immunological parameters in pigs.

Previous studies reported very low carry-over of dietary deoxynivalenol (DON) into eggs of laying hens. However, recent studies showed that DON is extensively metabolised to DON-3-sulphate (DON-3S) in chickens. We therefore hypothesised that DON-3S might also be a major DON metabolite in eggs of laying hens fed with DON contaminated diet. The aim of the work was to develop, validate and apply an LC-MS/MS based method for determination of DON, deepoxy-DON (DOM), DON-3S, and DOM-3-sulphate (DOM-3S) in freeze-dried eggs of laying hens. Laying hens were allocated to three treatment groups (negative control (NC); DON low (3.8 mg/kg DON in feed); DON high (7.5 mg/kg DON in feed)) and eggs were collected in the 5th, 7th and 10th week of the trial. DON-3S was identified as the major DON metabolite in eggs for the first time with average concentrations in fresh eggs

Fumonisin B1 (FB1), mainly produced by Fusarium verticillioides and Fusarium proliferatum, can be converted to the less toxic metabolite hydrolyzed FB1 (HFB1) by enzymatic degradation. The application of an FB1degrading enzyme as a feed additive is a strategy to reduce fumonisin exposure of animals. However, the difference between the effect of FB1 and HFB1 on porcine intestinal immunity is poorly documented. We investigated the toxic effects of FB1 and HFB1 exposure on porcine gut barrier function and intestinal immunity by using a co-culture model of intestinal porcine epithelial cells (IPEC-J2) and porcine peripheral blood mononuclear cells (PBMCs). First, we confirmed that Fusarium mycotoxin (deoxynivalenol; DON), in the presence of an endotoxin (lipopolysaccharide: LPS), disrupted gut permeability of IPEC-J2 and induced inflammatory response in the co-culture system. FB1 induced additional damage to gut barrier function and promoted pro-inflammatory responses in the presence of LPS and DON compared to only LPS/DON treatment. In the co-culture system, FB1/LPS/DON induced increased cell death of PBMCs and pro-inflammatory cytokines than LPS/DON treatment. In contrast, the application of HFB1 resulted in reduced levels of chemokines and pro-inflammatory cytokines together with marginal immune cell death compared to FB1/LPS/DON in the IPEC-J2/PBMC co-culture system. These findings suggest that FB1 aggravates LPS/DON-induced intestinal inflammation, and HFB1 showed less toxicity to immune response. Therefore, enzymatic degradation of FB1 to HFB1 could be an effective strategy to reduce intestinal inflammation in pigs.

In recent years, the deleterious effects attributed to mycotoxins, in particular on the intestine, faced increased attention and it was shown that deoxynivalenol (DON) causes adverse effects on gut health. In this context, it has been repeatedly reported that DON can alter the intestinal morphology, disrupt the intestinal barrier and reduce nutrient absorption. The underlying mechanism of a compromised intestinal barrier caused by DON in chickens has yet to be illustrated. Although, DON is rapidly absorbed from the upper parts of the small intestine, the effects on the large intestine cannot be excluded. Additionally, a damaging effect of DON on the gut epithelium might decrease the resistance of the gut against infectious agents. Consequently, the objectives of the present studies were: (1) to investigate the impact of DON on the epithelial paracellular permeability by demonstrating the mucosal to serosal flux of 14C-mannitol in the small and large intestine applying Ussing chambers and (2) to delineate the effects of DON on the colonization and translocation of Escherichia coli. Both parameters are well suited as potential indicators for gut barrier failure. For this, a total of 75 one-day-old Ross 308 broiler chickens were housed in floor pens on wood shavings with feed and water provided ad libitum. Birds were randomly allocated to three different groups (n = 25 with 5 replicates/group) and were fed for 5 weeks with either contaminated diets (5 or 10 mg DON/kg feed) or basal diets (control). Body weight (BW) and BW gain of birds in the group fed with 10 mg/kg DON were significantly lower than in group with 5 mg/kg DON and the control group. Moreover, the mannitol flux in jejunum and cecum was significantly (P

Mycotoxin intoxication is in general an acknowledged and tackled issue in animals. However, in several parts of the world, mycotoxicoses in humans still remain a relevant issue. The efficacy of two mycotoxin detoxifying animal feed additives, an aflatoxin bentonite clay binder and a fumonisin esterase, was investigated in a human child gut model, i.e. the in vitro Simulator of the Human Intestinal Microbial Ecosystem (SHIME®). Additionally, the effect of the detoxifiers on gut microbiota was examined in the SHIME. After an initial two weeks of system stabilisation, aflatoxin B1 (AFB1) and fumonisin B1 (FB1) were added to the SHIME diet during one week. Next, the two detoxifiers and mycotoxins were added to the system for an additional week. The AFB1, FB1, hydrolysed FB1 (HFB1), partially hydrolysed FB1a and FB1b concentrations were determined in SHIME samples using a validated ultra-performance liquid chromatography-tandem mass spectrometry method. The short-chain fatty acid (SCFA) concentrations were determined by a validated gas chromatography-mass spectrometry method. Colonic bacterial communities were analysed using metabarcoding, targeting the hypervariable V1-V3 regions of the 16S rRNA genes. The AFB1 and FB1 concentrations significantly decreased after the addition of the detoxifiers. Likewise, the concentration of HFB1 significantly increased. Concentrations of SCFAs remained generally stable throughout the experiment. No major changes in bacterial composition occurred during the experiment. The results demonstrate the promising effect of these detoxifiers in reducing AFB1 and FB1 concentrations in the human intestinal environment, without compromising the gastrointestinal microbiota.

Mycotoxins deoxynivalenol (DON) and zearalenone (ZEN) can negatively affect pig health. However, little is known about their effects on boar semen. We assessed the individual and combined effects of DON and ZEN on boar semen in vitro. In a pretrial, we determined the minimum dose (MiD) of each mycotoxin that induces a significant alteration of sperm progressive motility, as investigated using computer-assisted semen analysis (CASA). In the main trial, the individual and combined effects of each mycotoxin&rsquo, s MiD on sperm motility and kinetics (CASA analysis), morphology (SpermBlue staining), viability (calcein-propidium iodide staining), membrane functional status (hypoosmotic swelling test), and chromatin integrity (acridine orange staining) were analyzed. Pretrial results suggested a MiD of 50.6 &mu, M and 62.8 &mu, M for DON and ZEN, respectively. In the main trial, DON and ZEN administered at MiD significantly affected CASA parameters (e.g., increase of immotile spermatozoa, reduction of progressive motile spermatozoa), decreased sperm viability, and affected sperm morphology (head abnormalities) and membrane functional status. DON and ZEN showed less than additive effects on most parameters tested and a synergistic effect on viability and on two CASA parameters. In conclusion, DON and ZEN showed individual and combined toxic effects on boar semen in vitro.

The toxicokinetics (TK) of hydrolyzed fumonisin B1 (HFB1) were evaluated in 16 broiler chickens after being fed either a control or a fumonisins-contaminated diet (10.8 mg fumonisin B1, 3.3 mg B2 and 1.5 mg B3/kg feed) for two weeks, followed by a single oral (PO) or intravenous (IV) dose of 1.25 mg/kg bodyweight (BW) of HFB1. Fumonisin B1 (FB1), its partially hydrolyzed metabolites pHFB1a and pHFB1b, and fully hydrolyzed metabolite HFB1, were determined in chicken plasma using a validated ultra-performance liquid chromatography&ndash, tandem mass spectrometry method. None of the broiler chicken showed clinical symptoms of fumonisins (FBs) or HFB1 toxicity during the trial, nor was an aberration in body weight observed between the animals fed the FBs-contaminated diet and those fed the control diet. HFB1 was shown to follow a two-compartmental pharmacokinetic model with first order elimination in broiler chickens after IV administration. Toxicokinetic parameters of HFB1 demonstrated a total body clearance of 16.39 L/kg&middot, h and an intercompartmental flow of 8.34 L/kg&middot, h. Low levels of FB1 and traces of pHFB1b were found in plasma of chickens fed the FBs-contaminated diet. Due to plasma concentrations being under the limit of quantification (LOQ) after oral administration of HFB1, no toxicokinetic modelling could be performed in broiler chickens after oral administration of HFB1. Moreover, no phase II metabolites, nor N-acyl-metabolites of HFB1 could be detected in this study.

Mycotoxins, secondary metabolites produced by moulds, are responsible for causing significant economic losses due to spoilage of agricultural products but also due to direct or indirect health impact on livestock upon ingestion of mycotoxin contaminated feedstuffs. Aquaculture farmed species are not an exception and studies reporting mycotoxin‐related issues in the aquaculture industry have been increasing. However, our understanding on the prevalence and impact of mycotoxins in the aquaculture sector is still lower compared to the terrestrial livestock sector. Consequently, regulatory limits and guidance values have been defined based on the studies on terrestrial farm animals. The aim of this review is to compile and critically assess mycotoxin occurrence and co‐occurrence in aquaculture finished feeds, and understand the risk of mycotoxin carry‐over in aquaculture seafood products. Furthermore, we aim with this review to raise awareness to the scientific community, the regulatory authorities and the aquaculture industry to the need for specific aquaculture mycotoxin maximum concentration levels for both aquaculture feeds and foods.

Little is known about the effects of commonly found levels of Fusarium mycotoxins on the performance, metabolism, and immunity of dairy cattle. We investigated the effects of regular contamination levels, meaning contamination levels that can be commonly detected in dairy feeds, of deoxynivalenol (DON) and fumonisins (FB) in total mixed ration (TMR) on the performance, diet digestibility, milk quality, and plasma liver enzymes in dairy cows. This trial examined 12 lactating Holstein dairy cows using a 3-period × 3-treatment Latin square design. The experimental period was 21 d of mycotoxin exposure followed by 14 d of washout. During treatment periods, cows received one of 3 diets: (1) CTR (control) diet of TMR contaminated with 340.5 µg of DON/kg of dry matter (DM) and 127.9 µg FB/kg of DM; (2) MTX diet of TMR contaminated with Fusarium mycotoxins at levels higher than CTR but below US and European Union guidelines (i.e., 733.0 µg of DON/kg of DM and 994.4 µg of FB/kg of DM); or (3) MDP diet, which was MTX diet supplemented with a mycotoxin deactivator product (i.e., 897.3 µg of DON/kg of DM and 1,247.1 µg of FB/kg of DM; Mycofix, 35 g/animal per day). During washout, all animals were fed the same CTR diet. Body weight, body condition score, DM intake, dietary nutrient digestibility, milk production, milk composition and rennet coagulation properties, somatic cell count, blood serum chemistry, hematology, serum immunoglobulin concentrations, and expression of multiple genes in circulating leucocytes were measured. Milk production was significantly greater in the CTR group (37.73 kg/d) than in the MTX (36.39 kg/d) and the MDP (36.55 kg/d) groups. Curd firmness and curd firming time were negatively affected by the MTX diet compared with the other 2 diets. Furthermore, DM and neutral detergent fiber digestibility were lower after the MTX diet than after the CTR diet (67.3 vs. 71.0% and 42.8 vs. 52.3%). The MDP diet had the highest digestibility coefficients for DM (72.4%) and neutral detergent fiber (53.6%) compared with the other 2 diets. The activities of plasma liver transaminases were higher after the MTX diet than after the CTR and MDP diets. Compared with the CTR diet, the MTX diet led to slightly lower expression of genes related to immune and inflammatory functions, indicating that Fusarium mycotoxins had an immunosuppressive effect. Our results indicated that feed contaminated with regular levels of Fusarium mycotoxins adversely affected the performance, milk quality, diet digestibility, metabolic variables, and immunity of dairy cows, and that supplementation with mycotoxin deactivator product counteracted most of these negative effects.

Deoxynivalenol (DON) is one of the most prevalent Fusarium mycotoxins in grain and can cause economic losses in pig farming due to reduced feed consumption and lower weight gains. Biodetoxification of mycotoxins using bacterial strains has been a focus of research for many years. However, only a few in vivo studies have been conducted on the effectiveness of microbial detoxification of fusariotoxins. This study was therefore aimed at investigating the effect of a feed additive containing the bacterial strain Coriobacteriaceum DSM 11798 (the active ingredient in Biomin® BBSH 797) on growth performance and blood parameters, as well as uptake and metabolism of DON, in growing pigs. Forty-eight crossbred (Landrace-Yorkshire/Duroc-Duroc) weaning pigs were fed with pelleted feed made from naturally contaminated oats, with DON at four concentration levels: (1) control diet (DON < 0.2 mg kg−1), (2) low-contaminated diet (DON = 0.92 mg kg−1), (3) medium-contaminated diet (DON = 2.2 mg kg−1) and (4) high-contaminated diet (DON = 5.0 mg kg−1) and equivalent diets containing DSM 11798 as feed additive. During the first 7 days of exposure, pigs in the highest-dose group showed a 20–28% reduction in feed intake and a 24–34% reduction in weight gain compared with pigs in the control and low-dose groups. These differences were levelled out by study completion. Towards the end of the experiment, dose-dependent reductions in serum albumin, globulin and total serum protein were noted in the groups fed with DON-contaminated feed compared with the controls. The addition of DSM 11798 had no effect on the DON-related clinical effects or on the plasma concentrations of DON. The ineffectiveness of the feed additive in the present study could be a consequence of its use in pelleted feed, which might have hindered its rapid release, accessibility or detoxification efficiency in the pig’s gastrointestinal tract.

The awareness of mycotoxin-related issues in aquaculture industry has grown as manufacturers and producers realise the importance of mycotoxins, beyond aflatoxin, and their potential to impact production. During a period of one year (January 2016 – December 2016), 175 samples of different plant proteins, aquaculture/fishery by-products and finished aquaculture feeds, were analysed. Samples were tested for aflatoxins (aflatoxin B1, B2, G1 and G2); zearalenone; type B trichothecenes (deoxynivalenol (DON); nivalenol; 3-acetyldeoxynivalenol; 15-acetyldeoxynivalenol and fusarenon X-glucoside); fumonisins (fumonisin B1, B2 and B3); type A trichothecenes (T-2 and HT-2 toxin; diacetoxyscirpenol and neosolaniol); and ochratoxin A. All samples were sourced in Southeast Asia. Within the analysed samples, 4% were free of detectable mycotoxins. 8% of the samples had one mycotoxin and 88% of the samples were contaminated with more than one mycotoxin. The current work show similarity to previous studies, that mycotoxin occurrence in plant-meals, and consequently in finished feeds, is quite variable and confirms that aflatoxin is not the main mycotoxin in aquafeeds. Mycotoxin occurrence in fish and shrimp feeds in the present samples were higher than reported in previous studies for the same region. In samples of shrimp feed with DON detected, the mean average DON level was 882 μg/kg with a maximum level of 2,287 μg/kg. These values are within the reported sensitivity levels of white leg shrimp (Litopenaeus vannamei). Despite the limited number and short period of sampling, this inventory study helps address a knowledge gap of mycotoxin occurrence in feeds specifically destined for the aquaculture industry.

The objective of the present study was to demonstrate the efficiency of the decontamination process applied to deoxynivalenol (DON)-contaminated maize by sodium sulphite (Na2SO3) treatment in vivo. Additionally, in vitro characterisation of the toxicity of the DON sulphonates (DONS 1, 2 and 3 denote structurally different forms), the resulting DON metabolites, on peripheral blood mononuclear cells (PBMC) should substantiate the inactivation of DON. In a piglet experiment, both DON-contaminated maize and -uncontaminated control maize either untreated (DON-, CON-) or Na2SO3-treated (DON+, CON+) were mixed into feed and fed for 42 d starting from weaning. The results showed that feed intake and daily weight gain of animals fed DON- were significantly lower compared to animals fed CON- and CON+, whereas group DON+ reached the control level or even exceeded it. The feed-to-gain ratio was unaffected (p = 0.45). Furthermore, DON concentrations in plasma markedly reflected the diets' DON concentrations. These were < 0.1, < 0.1, 5.4 and 0.8 mg/kg feed for CON-, CON+, DON- and DON+, and amounted to 0.3, 0.4, 33.0 and 9.3 ng/ml in plasma, respectively. Whereas DONS 2 and 3 were detected in the DON+ diet, only DONS 2 was recovered in plasma. Regarding the toxicity of DONS, no or much lower toxicity was found compared to DON. DONS 1 and Na2SO3 did not affect the viability of PBMC. At 32.71μM DONS2 the viability was reduced by 50% and thus this compound was less toxic than DON by a factor of 73. Consequently, wet preservation of maize with Na2SO3 was an effective tool to avoid the adverse effects of DON on performance of piglets.

Concern about mycotoxins in aquaculture has been growing, partly due to the gradual replacement of animal-derived proteins, such as fish meal, by plant sources. Over a period of one year, 2,176 samples of different plant protein sources and 25 samples of finished aquaculture feeds were analysed. Samples were tested for aflatoxins (AF; sum of aflatoxin B1, B2, G1 and G2), zearalenone (ZEA), deoxynivalenol (DON), fumonisins (FB; sum of fumonisin B1 and B2), T-2 toxin and ochratoxin A (full toxin screen). The following plant-based meals were analysed: soybean meal (SBM), wheat (WH), wheat bran (WB), maize (C), corn gluten meal (CGM), cottonseed meal (CSM), rapeseed/canola meal (R/CM) and rice bran (RB). The plant raw materials and aquaculture finished feeds were obtained from Asia and Europe. Finished feed samples from Asia were acquired from Vietnam, Indonesia and Myanmar, while from Europe they were sampled from Denmark, Austria, the Netherlands and Germany. Mycotoxins were found in most of the commodities and finished feeds analysed, showing that mycotoxins represent a risk for the development of the aquaculture sector. Generally, in Asian samples we observed that SBM, WH, WB, C, CGM, R/CM and RB were mostly contaminated with Fusarium mycotoxins (ZEA, DON and FB). CSM was contaminated primarily by AF and Fusarium toxins (ZEA and DON) at lower concentrations. European samples were contaminated mainly by Fusarium mycotoxins. The co-occurrence of mycotoxins in all commodities was high, raising the probability of co-occurrence in finished feeds. An accumulation of mycotoxins on the processed plant-based ingredients (e.g. CGM and WB) was observed when compared to the respective whole grains (C and WH, respectively). Compared to results obtained in 2014, finished feeds presented lower contamination levels, but the co-occurrence risk increased.

A 10-week feeding experiment was carried out examining the effects of deoxynivalenol (DON)-contaminated maize treated with different sodium sulphite (SoS) concentrations on performance, health and DON-plasma concentrations in fattening pigs. Two maize batches were used: background-contaminated (CON, 0.73 mg/kg maize) and Fusarium-toxin contaminated (DON, 44.45 mg/kg maize) maize. Both were wet preserved at 20% moisture content, with one of three (0.0, 2.5, 5.0 g/kg maize) sodium sulphite concentrations and propionic acid (15%). Each maize batch was then mixed into a barley-wheat-based diet at a proportion of 10%, resulting in the following 6 feeding groups: CON− (CON + 0.0 g SoS/kg maize), CON2.5 (CON + 2.5 g SoS/kg maize), CON5.0 (CON + 5.0 g SoS/kg maize), DON- (DON + 0.0 g SoS/kg maize), DON2.5 (DON + 2.5 g SoS/kg maize) and DON5.0 (DON + 5.0 g SoS/kg maize). Dietary DON concentration was reduced by ~ 36% in group DON2.5 and ~ 63% in group DON5.0. There was no impact on ZEN concentration in the diets due to SoS treatment. Pigs receiving diet DON- showed markedly lower feed intake (FI) compared to those fed the control diets. With SoS-treatment of maize, FI of pigs fed the DON diet (DON5.0: 3.35 kg/d) were comparable to that control (CON−: 3.30 kg/day), and these effects were also reflected in live weight gain. There were some effects of SoS, DON or their interaction on serum urea, cholesterol and albumin, but always within the physiological range and thus likely negligible. SoS wet preservation of Fusarium-toxin contaminated maize successfully detoxified DON to its innocuous sulfonates, thus restoring impaired performance in fatteners.

The mycotoxin zearalenone (ZEN) is a frequent contaminant of animal feed and is well known for its estrogenic effects in animals. Cattle are considered less sensitive to ZEN than pigs. However, ZEN has previously been shown to be converted to the highly estrogenic metabolite &alpha, zearalenol (&alpha, ZEL) in rumen fluid in vitro. Here, we investigate the metabolism of ZEN in the reticulorumen of dairy cows. To this end, rumen-fistulated non-lactating Holstein Friesian cows (n = 4) received a one-time oral dose of ZEN (5 mg ZEN in 500 g concentrate feed) and the concentrations of ZEN and ZEN metabolites were measured in free rumen liquid from three reticulorumen locations (reticulum, ventral sac and dorsal mat layer) during a 34-h period. In all three locations, &alpha, ZEL was the predominant ZEN metabolite and &beta, zearalenol (&beta, ZEL) was detected in lower concentrations. ZEN, &alpha, ZEL and &beta, ZEL were eliminated from the ventral sac and reticulum within 34 h, yet low concentrations of ZEN and &alpha, ZEL were still detected in the dorsal mat 34 h after ZEN administration. In a second step, we investigated the efficacy of the enzyme zearalenone hydrolase ZenA (EC 3.1.1.-, commercial name ZENzyme&reg, BIOMIN Holding GmbH, Getzersdorf, Austria) to degrade ZEN to the non-estrogenic metabolite hydrolyzed zearalenone (HZEN) in the reticulorumen in vitro and in vivo. ZenA showed a high ZEN-degrading activity in rumen fluid in vitro. When ZenA was added to ZEN-contaminated concentrate fed to rumen-fistulated cows (n = 4), concentrations of ZEN, &alpha, ZEL were significantly reduced in all three reticulorumen compartments compared to administration of ZEN-contaminated concentrate without ZenA. Upon ZenA administration, degradation products HZEN and decarboxylated HZEN were detected in the reticulorumen. In conclusion, endogenous metabolization of ZEN in the reticulorumen increases its estrogenic potency due to the formation of &alpha, ZEL. Our results suggest that application of zearalenone hydrolase ZenA as a feed additive may be a promising strategy to counteract estrogenic effects of ZEN in cattle.

Ruminal microbiota of cattle are not able to detoxify all mycotoxins. In addition, detoxification can be hampered by adverse ruminal conditions (e.g., low ruminal pH). Hence, in the cattle husbandry, mycotoxin binders and modifiers could be used to prevent animal exposure to mycotoxins. In this study, an in vitro rumen model, including feed matrix, was established as first screening tool to test the efficacy of five products claiming to detoxify mycotoxins. The detoxifiers had different modes of action: (a) binding (three products), (b) enzymatic detoxification of zearalenone (ZEN, one product, ZenA), and (c) bacterial transformation of trichothecenes (one product, BBSH 797). For the mycotoxin binders, the binding to the mycotoxins enniatin B (ENN B), roquefortine C (ROQ-C), mycophenolic acid (MPA), deoxynivalenol (DON), nivalenol (NIV), and zearalenone (ZEN) were tested at a dose recommended by the manufacturers. The in vitro model demonstrated that all binders adsorbed ENN B to a certain extent, while only one of the binders also partially adsorbed ROQ-C. The binders did not change the concentrations of the other mycotoxins in the ruminal fluid. The enzyme ZenA detoxified ZEN very quickly and prevented the formation of the more toxic metabolite &alpha, zearalenol (&alpha, ZEL), both at normal (6.8) and low ruminal pH (5.8). The addition of BBSH 797 enhanced detoxification of DON and NIV, both at normal and low ruminal pH. The in vitro rumen model demonstrated that the addition of ZenA seems to be a very promising strategy to prevent estrogenic effects of ZEN contaminated feed, and BBSH 797 is efficient in the detoxification of trichothecenes.

Fumonisin B

We investigated the effects of feeding sodium sulfite (SoS) treated uncontaminated and Fusarium contaminated maize in a porcine lipopolysaccharide (LPS) challenge model. Eighty piglets (7.59 &plusmn, 0.92 kg body weight [BW]) were equally assigned to one of four experimental diets containing 10% maize, either uncontaminated and untreated (CON&minus, 0.09 mg deoxynivalenol [DON]/kg diet) or uncontaminated and SoS-treated (CON+, wet-preserved with 5 g SoS/kg maize, 0.05 mg DON/kg diet), or prepared with 10% of a Fusarium contaminated maize containing mainly deoxynivalenol (DON), either contaminated and untreated (FUS&minus, 5.36 mg DON/kg diet), or contaminated and SoS-treated (FUS+, wet-preserved with 5 g SoS/kg maize, 0.83 mg DON/kg diet). At day 42 of experiment, ten pigs of each group were injected intraperitoneally with either 7.5 &micro, g LPS/kg BW or placebo (0.9% NaCl). At 120 min after injection, blood samples were collected to analyse TNF-&alpha, hematological profile, clinical biochemistry as well as the redox status. A significant increase in body temperature and cytokine TNF-&alpha, concentration was observed in the LPS-injected piglets. Results for hematology, clinical chemistry and redox status indicate no effects of SoS treatment, with exception of neutrophil counts being significantly more pronounced after feeding the SoS treated FUS maize. In conclusion, SoS treatment of maize did not modulate the LPS-induced acute inflammation.

The impact of deoxynivalenol (DON) on rainbow trout, Oncorhynchus mykiss, is mainly characterised by impaired growth performance and reduced feed intake, usually with the total absence of any visible clinical signs. Despite the high concentrations of DON in the present study (up to 11,412 ± 1141 μg kg−1), no clinical signs (except anorexia at the higher DON dosage) were observed, which confirms the difficulties of diagnosing DON ingestion. Compared to the control group, the proteolytic enzyme activities (pepsin, trypsin and chymotrypsin) in trout were altered by DON ingestion. However, it was not clear if the observed impact on digestive enzymes was due to the direct action of DON, or a consequence of the lower feed intake determined for DON-treated animals. The impact of DON on the abundance of specific measured mRNA transcripts was unexpected with higher expression levels for insulin-like growth factors, igf1 and igf2, which are directly related to elevated insulin levels in plasma. This can also in part be influenced by the trypsin activity and by npy, given its higher mRNA expression levels. The apparent digestibility of dry matter, protein and energy was not affected by dietary levels of DON, however, nutrient retention, protein, fat and energy retention were significantly affected in animals fed DON. Adenylate cyclase-activating polypeptide (PACAP) expression seems to play an important role in controlling feed intake in DON fed trout. In the present study, we have shown for the first time that DON is metabolized to DON-3-sulfate in trout. DON-3-sulfate is much less toxic than DON, which helps to explain the lack of clinical signs in fish fed DON. Being a novel metabolite identified in trout makes it a potential biomarker of DON exposure. Suppression of appetite due to DON contamination in feeds might be a defense mechanism in order to decrease the exposure of the animal to DON, therefore reducing the potential negative impacts of DON., The present work was performed under the WiseFeed project (European Union's Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement No 691150) being Biomin Holding GmbH, a partner of the project, represented by Rui A. Gonçalves in his secondment to Instituto de Ciencias Marinas de Andalucía, Consejo Superior de Investigaciones Científicas (ICMAN-CSIC), Puerto Real, Cádiz, Spain.

We examined the toxicokinetics of fumonisin B1 (FB1) and its main metabolites after single dose application intravenously (iv) of 139 nmol FB1 or hydrolyzed FB1 (HFB1)/kg bodyweight (BW) in barrows (BW: 34.4 kg ± 2.7 kg), as well as the toxicokinetics of FB1, FB2, FB3 and FB1 bioavailability from oral exposure (3425 nmol FB1/kg BW, on top of ration). Additionally, detoxification efficacy of FumD (240 U/kg feed; 3321 nmol FB1/kg BW), a fumonisin esterase, was examined for oral fumonisin application. Urine and feces were collected quantitatively and serum samples were taken over a period of 120 h. Serum toxicokinetics of FB1iv showed a short distribution half-life of 6 min followed by a longer elimination half-life of 36 min. After HFB1iv administration, serum clearance was three times higher compared to FB1iv group (5.6 and 1.8 L/kg/h respectively) which together with a 5-times higher volume of distribution indicates that HFB1 is more rapidly cleared from systemic circulation but distributed more extensively into the extravasal space than FB1. The bioavailability of FB1 in orally exposed pigs was 5.2% (incl. metabolites). Moreover, we found a significant reduction of FB1 bioavailability by 90% caused by the action of fumonisin esterase in the gastrointestinal tract, clearly demonstrating the efficacy of FumD.

A feeding experiment with piglets was performed to examine the efficacy of a wet preservation of Fusarium (FUS)-contaminated maize with sodium sulphite (SoS) based on deoxynivalenol (DON) and zearalenone (ZEN) residue levels in urine, bile and liquor and health traits of piglets. For this purpose, 80 castrated male piglets (7.57 ± 0.92 kg BW) were assigned to four treatment groups: CON- (control diet, with 0.09 mg DON and0.01 mg ZEN/kg diet), CON+ (diet CON-, wet-preserved with 5 g SoS/kg maize; containing 0.05 mg DON and0.01 mg ZEN/kg diet), FUS- (diet with mycotoxin-contaminated maize; containing 5.36 mg DON and 0.29 mg ZEN/kg diet), and FUS+ (diet FUS-, wet-preserved with 5 g SoS/kg maize; resulting in 0.83 mg DON and 0.27 mg ZEN/kg diet). After 42 d, 40 piglets (n = 10 per group) were sampled. A clear reduction of DON levels by approximately 75% was detected in all specimens of pigs fed diet FUS+. ZEN was detected in all urine, bile and liquor samples, while their metabolites were only detectable in urine and bile. Additionally, their concentrations were not influenced by SoS treatment. Among the health-related traits, feeding of FUS diets increased the total counts of leukocytes and segmented neutrophil granulocytes irrespective of SoS treatment. SoS treatment increased the total blood protein content slightly with a similar numerical trend in albumin concentration. These effects occurred at an obviously lower level in FUS-fed groups. Moreover, SoS treatment recovered the reduction of NO production induced by feeding diet FUS- indicating an effect on the redox level. As this effect only occurred in group FUS+, it is obviously related to the adverse effects of the Fusarium toxins. In conclusion, treatment of FUS-contaminated maize with SoS decreased the inner exposure with DON as indicated by the lower DON levels in various piglet specimens. However, health-related traits did not consistently reflect this decreased exposure.

Deoxynivalenol (DON), a Fusarium mycotoxin, is one of the most prevalent mycotoxins in aquafeeds. The toxicokinetics of DON are rarely studied in aquatic species. The present study used juvenile rainbow trout (Oncorhynchus mykiss) with a mean initial body weight of 7.72 ± 1.42 g in order to evaluate the pharmacokinetic behaviour and the metabolization of radiolabelled DON ([3H]-DON). In a first trial, 30 fish were tube-fed with four pellets containing a total of 125 ± 0.019 ng of [3H]-DON. At different sampling time points after feeding (1 h, 3 h, 6 h, 12 h or 24 h), the tissue distribution of the [3H]-DON was assessed by liquid scintillation counting. In a second trial, five fish were tube-fed four pellets containing a total of 663 ng of unlabelled-DON. Twenty-four hours after feeding, metabolites of DON excreted into the water were analysed by LC-MS/MS. [3H]-DON was detected in fish liver one hour after tube-feeding, indicating a rapid absorption of DON. In the first hour, [3H]-DON achieved its maximum in the gastro-intestinal tract (GIT) (20.56 ± 8.30 ng). However, 6.19 ± 0.83 ng of [3H]-DON was also detected in the water at this sampling time point. The fast excretion of [3H]-DON (above the average gastric emptying time of trout) might be related to its high-water solubility and consequent excretion with the fluid phase of the chyme. The amount of [3H]-DON in the GIT was stable during the first six hours. Such long transit time of DON through the GIT increases the potential for damage and absorption. The period between six and twelve hours seems to be the turning point in terms of DON excretion. Twelve hours after tube-feeding, the trout excreted 50.71 ± 22.17% of the tube-fed DON amount into water, while at the previous sampling time point (six hours) only 11.03 ± 6.09% were detected. These data suggest that an effective method for gastrointestinal DON detoxification in trout requires a period of action lower than six hours. In the present trial, no DON metabolites were detected in water.

Supplementation of animal feeds with mycotoxin binders is routinely used to mitigate mycotoxin challenges. Adsorption efficiency of a binder is dependent on the structure and chemical properties of the mycotoxin, binder, and the complex gastrointestinal environment. Limited adsorption of non-polar mycotoxins has driven innovation of mitigation strategies including biotransformation or detoxification of mycotoxins by microorganisms and enzymatic activity. The current objective was to measure the efficacy of Biofix® Plus PRO (BPL) within a molasses-based liquid feed supplement to detoxify zearalenone (ZEN) and deoxynivalenol (DON) in an in vitro rumen batch culture system. Twelve reactors were filled with 1 L of liquid (50% rumen fluid, 30% reverse osmosis water, 20% synthetic saliva) and 10 g of feed (43% chopped hay, 57% corn-soy-based concentrate). Three treatments were tested in quadruplicate: Control (CON; liquid mixture only), Mycotoxin (MTX; CON + mycotoxin culture material to achieve concentration of 100 ppb ZEN and 1000 ppb DON), and BPL (MTX + 20 g/L of molasses-based liquid feed supplement including 1.65% of BPL). Samples were taken at 0 (molasses-based liquid feed added to BPL reactors after 15 min of incubation), 1, 3, 6, and 24 h and analyzed for ZEN and DON by HPLC. Results were averaged by treatment. No ZEN or DON was detected at any time point in CON group. After 3 h, ZEN was reduced by 91% and DON reduced by 87% in BPL group, with no detectable mycotoxins by 6 h. After 24 h, ZEN and DON were reduced 35% and 28%, respectively in MTX group due to natural microbial activity of the rumen fluid. Overall, the addition of Biofix® Plus PRO reduced ZEN and DON levels under the rumen batch culture conditions.

Under moderate climatic conditions, deoxynivalenol (DON) contamination occurs frequently on cereals. Detoxification measures are required to avoid adverse effects on farm animals. In the present study, a wet preservation method with sodium sulfite (Na2SO3) and propionic acid was tested to titrate the optimum Na2SO3-dose for maximum DON reduction of contaminated maize kernels and meal and to examine the interaction between dose and moisture content in dependence on the preservation duration. The DON concentration decreased with increasing amounts of supplemented Na2SO3 and with increasing duration of the preservation period in a bi-exponential fashion. Additionally, the feed structure and moisture content had a significant influence on the decontaminating effect. Variants with 30% moisture content favored higher DON reduction rates compared to 14% moisture, but especially at low moisture contents, DON reduction was more pronounced in maize kernels than in maize meal. In addition to the decrease of DON, a concomitant formation of three different DON sulfonates was observed which differed in their formation pattern over the time course of preservation. The overall results and statistical analysis clarified that Na2SO3 addition of 10 g/kg maize at 30% moisture for eight days was necessary to obtain a complete DON reduction.