Fumonisins alone or mixed with other fusariotoxins increase the C22-24:C16 sphingolipid ratios in chicken livers, while deoxynivalenol and zearalenone have no effect.

Poultry feed is often contaminated with fumonisins, deoxynivalenol, and zearalenone, which can result in oxidative damage, inflammation and change in lipid metabolism. Although sphingolipids play key roles in cells, only the effects of fumonisins on the sphingolipidome are well-documented. In chickens, fumonisins have been shown to increase the sphinganine to sphingosine ratio and the C22-24:C16 sphingolipid ratio, which has been proposed as a new biomarker of toxicity. In this study, we used UHPLC-MSMS targeted analysis to measure the effect of fusariotoxins on sphingolipids in the livers of chickens fed with diets containing fusariotoxins administered individually and in combination, at the maximum levels recommended by the European Commission. Chickens were exposed from hatching until they reached 35 days of age. This study revealed for the first time that fumonisins, deoxynivalenol, and zearalenone alone and in combination have numerous effects on the sphingolipidome in chicken livers. A 30-50 % decrease in ceramide, dihydroceramide, sphingomyelin, dihydrosphingomyelin, monohexosylceramide and lactosylceramide measured at the class level was observed when fusariotoxins were administered alone, whereas a 30-100 % increase in dihydroceramide, sphingomyelin, dihydrosphingomyelin, and monohexosylceramide was observed when the fusariotoxins were administered in combination. For these different variables, strong significant interactions were observed between fumonisins and zearalenone and between fumonisins and deoxynivalenol, whereas interactions between deoxynivalenol and zearalenone were less frequent and less significant. Interestingly, an increase in the C22-24:C16 ratio of ceramides, sphingomyelins, and monohexosylceramides was observed in chickens fed the diets containing fumonisins only, and this increase was close when the toxin was administered alone or in combination with deoxynivalenol and zearalenone. This effect mainly corresponded to a decrease in sphingolipids with a fatty acid chain length of 16 carbons, whereas C22-24 sphingolipids were unaffected or increased. In conclusion the C22-24:C16 ratio emerged as a specific biomarker, with variations dependent only on the presence of fumonisins.

Targeted sphingolipidomics indicates increased C22-C24:16 ratios of virtually all assayed classes in liver, kidney, and plasma of fumonisin-fed chickens.

The biological properties of sphinganine-(d18:0)-, sphingosine-(d18:1)-, deoxysphinganine-(m18: 0)-, deoxysphingosine-(m18:1)-, deoxymethylsphinganine-(m17:0)-, deoxymethylsphingosine-(m17:1)-, sphingadienine-(d18:2)-, and phytosphingosine-(t18:0)-sphingolipids have been reported to vary, but little is known about the effects of fumonisins, which are mycotoxins that inhibit ceramide synthase, on sphingolipids other than those containing d18:0 and d18:1. Thirty chickens divided into three groups received a control diet or a diet containing 14.6 mg FB1 + FB2/kg for 14 and 21 days. No effects on health or performance were observed, while the effects on sphingoid bases, ceramides, sphingomyelins, and glycosylceramides in liver, kidney, and plasma varied. The t18:0 forms were generally unaffected by fumonisins, while numerous effects were found for m18:0, m18:1, d18:2, and the corresponding ceramides, and these effects appeared to be similar to those observed for d18:0-, and d18:1-ceramides. Partial least square discriminant analysis showed that d18:1- and d18:0-sphingolipids are important variables for explaining the partitioning of chickens into different groups according to fumonisins feeding, while m17:1-, m18:0-, m18:1-, d18:2-, and t18:0-sphingolipids are not. Interestingly, the C22-C24:C16 ratios measured for each class of sphingolipid increased in fumonisin-fed chickens in the three assayed matrices, whereas the total amounts of the sphingolipid classes varied. The potential use of C22-C24:C16 ratios as biomarkers requires further study.

Zymosan-Induced Murine Peritonitis Is Associated with an Increased Sphingolipid Synthesis without Changing the Long to Very Long Chain Ceramide Ratio.

Sphingolipids are key molecules in inflammation and defense against pathogens. Their role in dectin-1/TLR2-mediated responses is, however, poorly understood. This study investigated the sphingolipidome in the peritoneal fluid, peritoneal cells, plasma, and spleens of mice after intraperitoneal injection of 0.1 mg zymosan/mouse or PBS as a control. Samples were collected at 2, 4, 8, and 16 h post-injection, using a total of 36 mice. Flow cytometry analysis of peritoneal cells and measurement of IL-6, IL-1ß, and TNF-α levels in the peritoneal lavages confirmed zymosan-induced peritonitis. The concentrations of sphingoid bases, dihydroceramides, ceramides, dihydrosphingomyelins, sphingomyelins, monohexosylceramides, and lactosylceramides were increased after zymosan administration, and the effects varied with the time and the matrix measured. The greatest changes occurred in peritoneal cells, followed by peritoneal fluid, at 8 h and 4 h post-injection, respectively. Analysis of the sphingolipidome suggests that zymosan increased the de novo synthesis of sphingolipids without change in the C14-C18:C20-C26 ceramide ratio. At 16 h post-injection, glycosylceramides remained higher in treated than in control mice. A minor effect of zymosan was observed in plasma, whereas sphinganine, dihydrosphingomyelins, and monohexosylceramides were significantly increased in the spleen 16 h post-injection. The consequences of the observed changes in the sphingolipidome remain to be established.

Targeted Sphingolipid Analysis in Heart, Gizzard, and Breast Muscle in Chickens Reveals Possible New Target Organs of Fumonisins.

Alteration of sphingolipid synthesis is a key event in fumonisins toxicity, but only limited data have been reported regarding the effects of fumonisins on the sphingolipidome. Recent studies in chickens found that the changes in sphingolipids in liver, kidney, lung, and brain differed greatly. This study aimed to determine the effects of fumonisins on sphingolipids in heart, gizzard, and breast muscle in chickens fed 20.8 mg FB1 + FB2/kg for 9 days. A significant increase in the sphinganine:sphingosine ratio due to an increase in sphinganine was observed in heart and gizzard. Dihydroceramides and ceramides increased in the hearts of chickens fed fumonisins, but decreased in the gizzard. The dihydrosphingomyelin, sphingomyelin, and glycosylceramide concentrations paralleled those of ceramides, although the effects were less pronounced. In the heart, sphingolipids with fatty acid chain lengths of 20 to 26 carbons were more affected than those with 14-16 carbons; this difference was not observed in the gizzard. Partial least squares-discriminant analysis on sphingolipids in the heart allowed chickens to be divided into two distinct groups according to their diet. The same was the case for the gizzard. Pearson coefficients of correlation among all the sphingolipids assayed revealed strong positive correlations in the hearts of chickens fed fumonisins compared to chickens fed a control diet, as well as compared to gizzard, irrespective of the diet fed. By contrast, no effect of fumonisins was observed on sphingolipids in breast muscle.

Targeted sphingolipid analysis in chickens suggests different mechanisms of fumonisin toxicity in kidney, lung, and brain.

Most of the toxic effects of fumonisins can be related to sphingolipid alteration, but there is little sphingolipidomic data in animals fed fumonisins in organs other than the liver. This study aimed to measure fumonisin B1 (FB1) in kidney, lung, and brain and determine its effects on sphingolipids. Thirty chickens divided into three groups received a diet containing 20.8 mg FB1+FB2/kg for 0, 4, or 9 days. FB1 increased in kidney from 1.7 to 5.6 nmol/kg and in lung from 0.5 to 1 nmol/kg at 4 and 9 days, respectively. No FB1 was detected in brain. In kidney, sphinganine increased, C14-C16 ceramides decreased, whereas C18-C26 ceramides increased. Most of the changes in dihydroceramides, dihydrodeoxyceramides, deoxyceramides sphingomyelins, dihydrosphingomyelins, and hexosylceramides paralleled those on ceramides. In lung, sphinganine was unaffected by fumonisins, whereas sphinganine-1-phosphate increased. Other major changes corresponded to decreases in glycosylceramides. In brain, sphinganine was unchanged, whereas deoxysphinganine, sphingosine, C14-C20 ceramides, and C14-C20 sphingomyelins increased. These results revealed that alterations in sphingolipids in kidney were close to those measured in liver and could correspond to inhibition of ceramide synthase 5 activity. In contrast, effects of fumonisins in lung and brain cannot be explained by inhibition of ceramide synthase.

Targeted Analysis of Sphingolipids in Turkeys Fed Fusariotoxins: First Evidence of Key Changes That Could Help Explain Their Relative Resistance to Fumonisin Toxicity.

The effects of fumonisins on sphingolipids in turkeys are unknown, except for the increased sphinganine to sphingosine ratio (Sa:So) used as a biomarker. Fumonisins fed at 20.2 mg/kg for 14 days were responsible for a 4.4 fold increase in the Sa:So ratio and a decrease of 33% and 36% in C14-C16 ceramides and C14-C16 sphingomyelins, respectively, whereas C18-C26 ceramides and C18-C26 sphingomyelins remained unaffected or were increased. Glucosyl- and lactosyl-ceramides paralleled the concentrations of ceramides. Fumonisins also increased dihydroceramides but had no effect on deoxysphinganine. A partial least squfares discriminant analysis revealed that all changes in sphingolipids were important in explaining the effect of fumonisins. Because deoxynivalenol and zearalenone are often found in feed, their effects on sphingolipids alone and in combination with fumonisins were investigated. Feeding 5.12 mg deoxynivalenol/kg reduced dihydroceramides in the liver. Zearalenone fed at 0.47 mg/kg had no effect on sphingolipids. When fusariotoxins were fed simultaneously, the effects on sphingolipids were similar to those observed in turkeys fed fumonisins alone. The concentration of fumonisin B1 in the liver of turkeys fed fumonisins was 0.06 µmol/kg. Changes in sphingolipid concentrations differed but were consistent with the IC50 of fumonisin B1 measured in mammals; these changes could explain the relative resistance of turkeys to fumonisins.

Strong Alterations in the Sphingolipid Profile of Chickens Fed a Dose of Fumonisins Considered Safe.

Fumonisins (FB) are mycotoxins known to exert most of their toxicity by blocking ceramide synthase, resulting in disruption of sphingolipid metabolism. Although the effects of FB on sphinganine (Sa) and sphingosine (So) are well documented in poultry, little information is available on their other effects on sphingolipids. The objective of this study was to analyze the effects of FB on the hepatic and plasma sphingolipidome in chickens. The first concern of this analysis was to clarify the effects of FB on hepatic sphingolipid levels, whose variations can lead to numerous toxic manifestations. The second was to specify the possible use of an alteration of the sphingolipidome as a biomarker of exposure to FB, in addition to the measurement of the Sa:So ratio already widely used. For this purpose, we developed an UHPLC MS/MS method that enabled the determination of 82 SL, including 10 internal standards, in chicken liver and plasma. The validated method was used to measure the effects of FB administered to chickens at a dose close to 20 mg FB1 + FB2/kg feed for 9 days. Significant alterations of sphingoid bases, ceramides, dihydroceramides, glycosylceramides, sphingomyelins and dihydrosphingomyelins were observed in the liver. In addition, significant increases in plasma sphinganine 1-phosphate, sphingosine 1-phosphate and sphingomyelins were observed in plasma. Interestingly, partial least-squares discriminant analysis of 11 SL in plasma made it possible to discriminate exposed chickens from control chickens, whereas analysis of Sa and So alone revealed no difference. In conclusion, our results show that the effects of FB in chickens are complex, and that SL profiling enables the detection of exposure to FB when Sa and So fail.

Fumonisin B1 Accumulates in Chicken Tissues over Time and This Accumulation Was Reduced by Feeding Algo-Clay.

The toxicokinetics of the food and feed contaminant Fumonisin B (FB) are characterized by low oral absorption and rapid plasma elimination. For these reasons, FB is not considered to accumulate in animals. However, recent studies in chicken and turkey showed that, in these species, the hepatic half-elimination time of fumonisin B1 (FB1) was several days, suggesting that FB1 may accumulate in the body. For the present study, 21-day-old chickens received a non-toxic dose of around 20 mg FB1 + FB2/kg of feed to investigate whether FB can accumulate in the body over time. Measurements taken after four and nine days of exposure revealed increased concentrations of sphinganine (Sa) and sphingosine (So) over time in the liver, but no sign of toxicity and no effect on performances were observed at this level of FB in feed. Measurements of FB in tissues showed that FB1 accumulated in chicken livers from four to nine days, with concentrations of 20.3 and 32.1 ng FB1/g observed, respectively, at these two exposure periods. Fumonisin B2 (FB2) also accumulated in the liver, from 0.79 ng/g at four days to 1.38 ng/g at nine days. Although the concentrations of FB found in the muscles was very low, an accumulation of FB1 over time was observed in this tissue, with concentrations of 0.036 and 0.072 ng FB1/g being measured after four and nine days of exposure, respectively. Feeding algo-clay to the chickens reduced the accumulation of FB1 in the liver and muscle by , approximately 40 and 50% on day nine, respectively. By contrast, only a weak non-significant effect was observed on day four. The decrease in the concentration of FB observed in tissues of chickens fed FB plus algo-clay on day nine was accompanied by a decrease in Sa and So contents in the liver compared to the levels of Sa and So measured in chickens fed FB alone. FB1 in the liver and Sa or So contents were correlated in liver tissue, confirming that both FB1 and Sa are suitable biomarkers of FB exposure in chickens. Further studies are necessary to determine whether FB can accumulate at higher levels in chicken tissues with an increase in the time of exposure and in the age of the animals.

Toxic Effects of Fumonisins, Deoxynivalenol and Zearalenone Alone and in Combination in Ducks Fed the Maximum EUTolerated Level.

Toxic effects among fumonisins B (FB), deoxynivalenol (DON) and zearalenone (ZEN) administered alone and combined were investigated in 84-day-old ducks during force-feeding. 75 male ducks, divided into five groups of 15 animals, received daily during the meal a capsule containing the desired among of toxin. Treated animals received dietary levels of toxins equivalent to 20 mg FB1+FB2/kg (FB), 5 mg DON/kg (DON), 0.5 mg ZEN/kg (ZEN) and 20, 5 and 0.5 mg/kg of FB, DON and ZEN (FBDONZEN), respectively. Control birds received capsules with no toxin. After 12 days, a decrease in body weight gain accompanied by an increase in the feed conversion ratio was observed in ducks exposed to FBDONZEN, whereas there was no effect on performances in ducks exposed to FB, DON and ZEN separately. No difference among groups was observed in relative organ weight, biochemistry, histopathology and several variables used to measure oxidative damage and testicular function. A sphinganine to sphingosine ratio of 0.32, 1.19 and 1.04, was measured in liver in controls and in ducks exposed to FB and FBDONZEN, respectively. Concentrations of FB1 in liver were 13.34 and 15.4 ng/g in ducks exposed to FB and FBDONZEN, respectively. Together ZEN and its metabolites were measured after enzymatic hydrolysis of the conjugated forms. Mean concentrations of α-zearalenol in liver were 0.82 and 0.54 ng/g in ducks exposed to ZEN and FBDONZEN, respectively. ß-zearalenol was 2.3-fold less abundant than α-zearalenol, whereas ZEN was only found in trace amounts. In conclusion, this study suggests that decreased performance may occur in ducks exposed to a combination of FB, DON and ZEN, but does not reveal any other interaction between mycotoxins in any of the other variables measured.

Mycotoxin and Gut Microbiota Interactions.

The interactions between mycotoxins and gut microbiota were discovered early in animals and explained part of the differences in susceptibility to mycotoxins among species. Isolation of microbes present in the gut responsible for biotransformation of mycotoxins into less toxic metabolites and for binding mycotoxins led to the development of probiotics, enzymes, and cell extracts that are used to prevent mycotoxin toxicity in animals. More recently, bioactivation of mycotoxins into toxic compounds, notably through the hydrolysis of masked mycotoxins, revealed that the health benefits of the effect of the gut microbiota on mycotoxins can vary strongly depending on the mycotoxin and the microbe concerned. Interactions between mycotoxins and gut microbiota can also be observed through the effect of mycotoxins on the gut microbiota. Changes of gut microbiota secondary to mycotoxin exposure may be the consequence of the antimicrobial properties of mycotoxins or the toxic effect of mycotoxins on epithelial and immune cells in the gut, and liberation of antimicrobial peptides by these cells. Whatever the mechanism involved, exposure to mycotoxins leads to changes in the gut microbiota composition at the phylum, genus, and species level. These changes can lead to disruption of the gut barrier function and bacterial translocation. Changes in the gut microbiota composition can also modulate the toxicity of toxic compounds, such as bacterial toxins and of mycotoxins themselves. A last consequence for health of the change in the gut microbiota secondary to exposure to mycotoxins is suspected through variations observed in the amount and composition of the volatile fatty acids and sphingolipids that are normally present in the digesta, and that can contribute to the occurrence of chronic diseases in human. The purpose of this work is to review what is known about mycotoxin and gut microbiota interactions, the mechanisms involved in these interactions, and their practical application, and to identify knowledge gaps and future research needs.

Zearalenone and Metabolites in Livers of Turkey Poults and Broiler Chickens Fed with Diets Containing Fusariotoxins.

Zearalenone (ZEN) and metabolites were measured in livers of turkeys and broilers fed a control diet free of mycotoxins, a diet that contained 0.5 mg/kg ZEN (ZEN diet), and a diet that contained 0.5, 5, and 20 mg/kg of ZEN, fumonisins, and deoxynivalenol, respectively (ZENDONFB diet). The feed was individually distributed to male Grade Maker turkeys from the 55th to the 70th day of age and to male Ross chickens from the 1st to the 35th day of age, without any signs of toxicity. Together, the free and conjugated forms of ZEN, α- and ß-zearalenols (ZOLs), zearalanone (ZAN), and α- and ß-zearalanols (ZALs) were measured by UHPLC-MS/MS with [13C18]-ZEN as an internal standard and immunoaffinity clean-up of samples. ZAN and ZALs were not detected. ZEN and ZOLs were mainly found in their conjugated forms. α-ZOL was the most abundant and was found at a mean concentration of 2.23 and 1.56 ng/g in turkeys and chickens, respectively. Consuming the ZENDONFB diet significantly increased the level of total metabolites in the livers of chickens. Furthermore, this increase was more pronounced for the free forms of α-ZOL than for the conjugated forms. An investigation of the presence of ZEN and metabolites in muscle with the methods validated for the liver failed to reveal any traces of these contaminants in this tissue. These results suggest that concomitant dietary exposure to deoxynivalenol (DON) and fumonisins (FB) may alter the metabolism and persistence of ZEN and its metabolites in the liver.

Toxicity of Fumonisins, Deoxynivalenol, and Zearalenone Alone and in Combination in Turkeys Fed with the Maximum European Union-Tolerated Level.

Surveys of mycotoxins worldwide have shown that deoxynivalenol (DON), fumonisins (FB), and zearalenone (ZON) are the most abundant Fusarium mycotoxins (FUS) in European poultry feed, in both the level and the frequency of contamination. Previous studies reported that a combination of FUS at concentrations that individually are not toxic may negatively affect animals. However, although toxic thresholds and regulatory guidelines exist for FUS, none account for the risk of multiple contamination, which is the most frequent. The aim of this study was to compare DON, FB, and ZON toxicity, alone and in combination, in male turkey poults. Ground cultured toxigenic Fusarium strains were incorporated in corn-soybean-based feed in five experimental diets: control diet, containing no mycotoxins, DON diet (5 mg DON/kg), FB diet (20 mg FB1 + FB2/ kg), ZON diet (0.5 mg ZON/kg), and DONFBZON diet (5, 20, and 0.5 mg/kg of DON, FB1 + FB2, and ZON, respectively). Seventy male Grade Maker turkeys were reared in individual cages on mycotoxin-free diets from 0 to 55 days of age. On the 55th day, the turkeys were weighed and divided into five groups each comprising 14 birds. Each group was fed one of the five experimental diets for a period of 14 days. On the 70th day of age, feed was withheld for 8 hr, at which time a blood sample was collected, and then all the turkeys were killed, autopsied, and different tissues sampled. The weight of the different organs, analyses of performance, biochemistry, histopathology, oxidative damage, and testis toxicity revealed no significant effects attributable to FUS. Measurement of sphingolipids in the liver revealed an increase in the sphinganine to sphingosine ratio in turkeys fed diets containing FB, but had no apparent consequences in terms of toxicity. Finally, only slight differences were found in some variables and the results of this study showed no interactions between DON, FB, and ZON. Taken together, results thus suggest that the maximum tolerated levels established for individual contamination by DON, FB, and ZON can also be considered safe in turkeys fed with combinations of these FUS for a period of 14 days.

Toxicidad de fumonisinas, deoxinivalenol y zearalenona solos y en combinación en pavos alimentados con el nivel máximo tolerado por la Unión Europea. Investigaciones sobre micotoxinas en todo el mundo han demostrado que el deoxinivalenol (DON), las fumonisinas (FB) y la zearalenona (ZON) son las micotoxinas de Fusarium (FUS) más abundantes en la alimentación avícola europea, tanto en el nivel como en la frecuencia de la contaminación. Estudios anteriores informaron que una combinación de micotoxinas de Fusarium a concentraciones que individualmente no son tóxicas puede afectar negativamente a los animales. Sin embargo, aunque existen umbrales tóxicos y pautas regulatorias para las micotoxinas de Fusarium, ninguno tiene en cuenta el riesgo de contaminación múltiple, que es lo más frecuente. El objetivo de este estudio fue comparar la toxicidad deoxinivalenol, fumonisinas, y zearalenona, solas o en combinación en pavos machos. Cepas toxigénicas de Fusarium cultivadas en suelos fueron incorporadas en alimentos a base de maíz y soya en cinco dietas experimentales: dieta de control, que no contiene micotoxinas, dieta DON (5 mg DON/kg), dieta FB (20 mg FB1+FB2/kg), dieta ZON (0.5 mg de ZON/kg) y dieta DONFBZON (5, 20 y 0.5 mg/kg de deoxinivalenol, fumonisinas 1 y 2 y zearalenona, respectivamente). Setenta pavos machos Grado Maker fueron criados en jaulas individuales con dietas libres de micotoxinas de 0 a 55 días de edad. En el día 55, los pavos se pesaron y se distribuyeron en cinco grupos, cada uno con 14 aves. Cada grupo fue alimentado con una de las cinco dietas experimentales durante un período de 14 días. En el día 70 de edad, el alimento se retuvo durante 8 horas, momento en el que se recolectó una muestra de sangre, y luego se sacrificaron todos los pavos, se les realizó la necropsia y se tomaron muestras de diferentes tejidos. El peso de los diferentes órganos, los análisis de rendimiento, la bioquímica, la histopatología, el daño oxidativo y la toxicidad en testículos no revelaron efectos significativos atribuibles a micotoxinas de Fusarium. La medición de esfingolípidos en el hígado reveló un aumento en la relación de esfinganina con relación a la esfingosina en pavos alimentados con dietas que contenían fumonisinas, pero no tuvo consecuencias aparentes en términos de toxicidad. Finalmente, solo se encontraron ligeras diferencias en algunas variables y los resultados de este estudio no mostraron interacciones entre deoxinivalenol, fumonisinas y zearalenona. Tomados en conjunto, los resultados sugieren que los niveles máximos tolerados establecidos para la contaminación individual por deoxinivalenol, fumonisinas y zearalenona también pueden considerarse seguros en pavos alimentados con combinaciones de estas micotoxinas de Fusarium durante un período de 14 días.

Unusual acute neonatal mortality and sow agalactia linked with ergot alkaloid contamination of feed.

BACKGROUND: An increase in the occurrence of ergot alkaloid contamination has been observed in Europe in recent years. The typical clinical signs of pig ergot poisoning are impaired growth, agalactia and, sometimes, gangrene. Opportunities for reporting exposure doses associated with clinical signs in animals under field conditions are rare. CASE PRESENTATION: In a farrow-to-finish pig farm with 160 sows, excessive acute neonatal mortality was reported in association with a loss of appetite and agalactia in sows. A herd examination was conducted and a high rate of piglet loss and agalactia in 13 sows out of the most affected batch of 20 were confirmed. Necropsy showed piglets with empty stomachs and intestines, with apparently normal mucosa. Gestating and lactating sow diet samples, as well as a wheat sample, were sent for analysis following feed mill inspection and a hypothesis of mycotoxin contamination of self-prepared feed. Liquid chromatography with mass spectrometry in tandem revealed an amount of total ergot alkaloids in all of the samples ranging from 3.49 mg/kg (gestating diet) to 8.06 mg/kg (lactating diet). The contaminated feed was removed and the situation returned to normal 3 weeks later (following batch of sows). CONCLUSION: In the present case, the exposure of sows to 3.49 mg/kg ergot alkaloid for 10 to 15 days before the end of gestation and to 8.06 mg/kg ergot alkaloid over 3 to 4 days at the beginning of lactation - corresponding to a content of 10,146 mg of sclerotia/kg in the wheat of the diets- led to agalactia in 13 of 20 sows in a batch and to a high neonatal mortality rates for all litters. No clinical signs associated with vasoconstrictive effects were observed.

Fumonisin B1, B2 and B3 in Muscle and Liver of Broiler Chickens and Turkey Poults Fed with Diets Containing Fusariotoxins at the EU Maximum Tolerable Level.

Although provisional maximum tolerable daily intake and recommended guidelines have been established for fumonisins (FB) in food, few data are available concerning levels of FB in edible animal tissues. Such data are of particular interest in avian species that can tolerate relatively high levels of fumonisins in their feed. Also, even if multiple contamination of animal feed by toxins produced by Fusarium is very frequent, little is known about the consequences of multiple contamination for FB levels in tissues. The aim of this study was to analyze the concentrations of FB in the muscle and liver of chickens and turkeys fed with FB alone and with FB combined with deoxynivalenol (DON), and with zearalenone (ZEN). Experimental diets were formulated by incorporating ground cultured toxigenic Fusarium strains in corn-soybean based feeds. Control diets were free of mycotoxins, FB diets contained 20 mg FB1+FB2/kg, and FBDONZEN diets contained 20, 5, and 0.5 mg/kg of FB1+FB2, DON, and ZEN, respectively. Animals were reared in individual cages with free access to water and feed. The feed was distributed to male Ross chickens from the 1st to the 35th day of age and to male Grade Maker turkeys from the 55th to the 70th day of age. On the last day of the study, the birds were starved for eight hours, killed, and autopsied for tissues sampling. No sign of toxicity was observed. A UHPLC-MS/MS method with isotopic dilution and immunoaffinity clean-up of samples has been developed for analysis of FB in muscle (n = 8 per diet) and liver (n = 8 per diet). Only traces of FB that were below the LOQ of 0.25 µg/kg were found in most of the samples of animals fed with the control diets. Mean concentrations of FB1, FB2, and FB3 in muscle were 17.5, 3.39, and 1.26 µg/kg, respectively, in chickens, and 5.77, 1.52, and 0.54 µg/kg in turkeys, respectively. In the liver, the respective FB1, FB2, and FB3 concentrations were 44.7, 2.61, and 0.79 µg/kg in chickens, and 41.47, 4.23, and 1.41 µg/kg, in turkeys. Cumulated level of FB1+FB2+FB3 in the highly contaminated samples were above 60 and 100 µg/kg in muscle and liver, respectively. The concentrations of FB in the tissues of animals fed the FBDONZEN diet did not greatly differ from the concentrations measured in animals fed the diet containing only FB.

Lack of Toxic Interaction Between Fusariotoxins in Broiler Chickens Fed throughout Their Life at the Highest Level Tolerated in the European Union.

Fusarium mycotoxins (FUS) occur frequently in poultry diets, and regulatory limits are laid down in several countries. However, the limits were established for exposure to a single mycotoxin, whereas multiple contamination is more realistic, and different studies have demonstrated that it is not possible to predict interactions between mycotoxins. The purpose of this study was thus to compare the toxic effect of deoxynivalenol (DON), fumonisins (FB) and zearalenone (ZON), alone and in combination on broiler chickens, at the maximum tolerated level established by the EU for poultry feed. Experimental corn-soybean diets incorporated ground cultured toxigenic Fusarium strains. One feed was formulated for chickens 0 to 10 days old and another for chickens 11 to 35 days old. The control diets were mycotoxin free, the DON diets contained 5 mg DON/kg, the FB diet contained 20 mg FB1 + FB2/kg, and the ZON diet contained 0.5mg ZON/kg. The DONFBZON diet contained 5, 20, and 0.5 mg/kg of DON, FB1 + FB2, and ZON, respectively. Diets were distributed ad libitum to 70 broilers (male Ross PM3) separated into five groups of 14 chickens each reared in individual cages from one to 35 days of age. On day 35, after a starvation period of 8 h, a blood sample was collected, and all the animals were killed and autopsied. No difference between groups that could be attributed to FUS was observed in performances, the relative weight of organs, biochemistry, histopathology, intestinal morphometry, variables of oxidative damage, and markers of testicle toxicity. A significant increase in sphinganine and in the sphinganine to sphingosine ratio was observed in broilers fed FB. Taken together, these results suggest that the regulatory guidelines established for single contamination of broiler chickens fed with DON, FB, and ZON can also be used in the case of multiple contamination with these toxins.

Worldwide Mycotoxins Exposure in Pig and Poultry Feed Formulations.

The purpose of this review is to present information about raw materials that can be used in pig and poultry diets and the factors responsible for variations in their mycotoxin contents. The levels of mycotoxins in pig and poultry feeds are calculated based on mycotoxin contamination levels of the raw materials with different diet formulations, to highlight the important role the stage of production and the raw materials used can have on mycotoxins levels in diets. Our analysis focuses on mycotoxins for which maximum tolerated levels or regulatory guidelines exist, and for which sufficient contamination data are available. Raw materials used in feed formulation vary considerably depending on the species of animal, and the stage of production. Mycotoxins are secondary fungal metabolites whose frequency and levels also vary considerably depending on the raw materials used and on the geographic location where they were produced. Although several reviews of existing data and of the literature on worldwide mycotoxin contamination of food and feed are available, the impact of the different raw materials used on feed formulation has not been widely studied.

Fusariotoxins in Avian Species: Toxicokinetics, Metabolism and Persistence in Tissues.

Fusariotoxins are mycotoxins produced by different species of the genus Fusarium whose occurrence and toxicity vary considerably. Despite the fact avian species are highly exposed to fusariotoxins, the avian species are considered as resistant to their toxic effects, partly because of low absorption and rapid elimination, thereby reducing the risk of persistence of residues in tissues destined for human consumption. This review focuses on the main fusariotoxins deoxynivalenol, T-2 and HT-2 toxins, zearalenone and fumonisin B1 and B2. The key parameters used in the toxicokinetic studies are presented along with the factors responsible for their variations. Then, each toxin is analyzed separately. Results of studies conducted with radiolabelled toxins are compared with the more recent data obtained with HPLC/MS-MS detection. The metabolic pathways of deoxynivalenol, T-2 toxin, and zearalenone are described, with attention paid to the differences among the avian species. Although no metabolite of fumonisins has been reported in avian species, some differences in toxicokinetics have been observed. All the data reviewed suggest that the toxicokinetics of fusariotoxins in avian species differs from those in mammals, and that variations among the avian species themselves should be assessed.

Ergot alkaloids produced by endophytic fungi of the genus Epichloë.

The development of fungal endophytes of the genus Epichloë in grasses results in the production of different groups of alkaloids, whose mechanism and biological spectrum of toxicity can differ considerably. Ergot alkaloids, when present in endophyte-infected tall fescue, are responsible for "fescue toxicosis" in livestock, whereas indole-diterpene alkaloids, when present in endophyte-infected ryegrass, are responsible for "ryegrass staggers". In contrast, peramine and loline alkaloids are deterrent and/or toxic to insects. Other toxic effects in livestock associated with the consumption of endophyte-infected grass that contain ergot alkaloids include the "sleepy grass" and "drunken horse grass" diseases. Although ergovaline is the main ergopeptine alkaloid produced in endophyte-infected tall fescue and is recognized as responsible for fescue toxicosis, a number of questions still exist concerning the profile of alkaloid production in tall fescue and the worldwide distribution of tall fescue toxicosis. The purpose of this review is to present ergot alkaloids produced in endophyte-infected grass, the factors of variation of their level in plants, and the diseases observed in the mammalian species as relate to the profiles of alkaloid production. In the final section, interactions between ergot alkaloids and drug-metabolizing enzymes are presented as mechanisms that could contribute to toxicity.

Ergovaline and lolitrem B concentrations in perennial ryegrass in field culture in southern France: distribution in the plant and impact of climatic factors.

Perennial ryegrass (Lolium perenne) infected by Epichloë festucae var. lolii contains alkaloids that are responsible for toxicosis in several countries, but few cases are reported in Europe. Lolitrem B is generally the most abundant alkaloid and is recognized to be responsible for livestock staggers, whereas ergovaline is less frequently documented in perennial ryegrass. Lolitrem B and ergovaline were monitored over a three-year period in endophyte-infected perennial ryegrass 'Samson' sown in southern France. Alkaloid concentrations were strongly influenced by the stage of maturity of the plant; maximum concentrations were always measured at the fully ripe stage. Over the three years of analysis, variations in lolitrem B in the whole plant at the fully ripe stage were low (from 1296 to maximum 1871 µg/kg dry matter), whereas ergovaline varied considerably (from 526 to 2322 µg/kg dry matter), suggesting that abiotic factors play a key role in determining ergovaline levels in endophyte-infected perennial ryegrass.

Endophyte infection of tall fescue and the impact of climatic factors on ergovaline concentrations in field crops cultivated in southern France.

Tall fescue (Lolium arundinaceum) infected by Epichloe coenophiala contains ergot alkaloids responsible for fescue toxicosis in Australia, New Zealand, and the United States, with only a few cases occurring in Europe. The detection of Epichloe in 166 L. arundinaceum collected in southern France revealed that 60% were infected, 51% being high ergovaline producers. The ergovaline level in endophyte-infected tall fescue Kentucky 31 was monitored during 3 years in various parts of the plant. Maturation of plants, recorded according to the BBCH scale, appeared to be the main factor for estimating the risk of toxicity. Ergovaline levels of ≥300 µg/kg dry matter were obtained at the end of spring, the beginning of autumn, and mid-winter. Positive correlation between ergovaline level and cumulative degree-d was observed, whereas rainfall had no effect. These results suggest that the lack of fescue toxicosis observed in France cannot be explained by the lack of ergovaline in tall fescue.