Effects of a deoxynivalenol contaminated diet on feed intake and health status in horses.

No description available.

F. graminearum est l’une des espèces prédominantes en tant que pathogène des céréales en Europe et dans les régions tempérées du monde. Cette moisissure appartient au genre Fusarium qui est l’un des genres qui produit le plus de métabolites secondaires, dont de nombreuses mycotoxines. Cependant, les connaissances sur son génome laissent présager que F. graminearum produit une très grande variété de métabolites qui sont encore inconnus. C’est pourquoi l’objectif de cette thèse a été de caractériser le métabolome de cette moisissure en identifiant de nouveaux métabolites ainsi qu’en étudiant leur toxicité. Pour cela, une méthode analytique combinant la spectrométrie de masse de haute résolution et le marquage des métabolites secondaires avec des isotopes stables a été développée. Celle‐ci à tout d’abord était validée avec le métabolome bien connu d’Aspergillus fumigatus avant d’être appliquée au métabolome de F. graminearum. Cette méthode a permis d’identifier 37 nouveaux métabolites, dont les fusaristatines A, C et D qui on été détectées pour la première fois chez F. graminearum. Le deoxynivalenol (DON) est une mycotoxine majeure produite par F. graminearum, c’est pourquoi sa toxicité a été comparée in vivo sur un modèle porcin à celle d’un mélange de fusariotoxines. Les résultats de cette étude ont montré une toxicité bien différente du mélange par rapport au DON seul, que ce soit par rapport à la performance des animaux ou à leur réponse immunitaire. Ceci suggère que certaines des fusariotoxines présentes dans le mélange auraient des répercutions biologiques et donc que leur toxicité reste encore à être élucidée. De plus, l’importance de la caractérisation de la toxicité du métabolome de F. graminearum a été soulignée par les résultats obtenus dans une seconde étude, qui ont révélés une possible implication du DON dans les maladies inflammatoires chroniques de l’intestin par l’intermédiaire des lymphocytes Th17. / F. graminearum is one of the most predominant cereal pathogen in Europe and temperate regions of the world. It belongs to the genera Fusarium, which is one of the most important in terms of the production of secondary metabolites, among which stand several mycotoxins. However, recent advances on the study of its genome suggest that only a small fraction of the secondary metabolites produced by F. graminearum have been identified so far. Therefore, the main objective of this thesis was to characterize the metabolome of this fungus by identifying new metabolites and by analyzing their toxicity. First, an analytical method combining high resolution mass spectrometry and isotopically labeled secondary metabolites was developed. The method was validated with the well known metabolome of Aspergillus fumigatus before it was applied to F. graminearum. As a result, 37 new metabolites were identified such as fusaristatin A, C and D, which were detected for the first time in the metabolome of F. graminearum. Deoxynivalenol (DON) is one of the major mycotoxins produced by this fungus. Therefore, its toxicity was compared to that of a blend of fusariotoxins in vivo in a pig model. The results of this study showed different effects of the blend compared to DON alone on animal performances and immune response. This suggests that some of the fusariotoxins present in the blend could be biologically active and their toxicity should be further investigated. Toxicity of the metabolome of F. graminearum has also been highlighted by the results of a second study which showed a possible implication of DON on the appearance of inflammatory bowel diseases by the action of Th17 lymphocytes.

Effects of a deoxynivalenol contaminated diet on feed intake and health status in horses.

No description available.

Master of Science / Department of Animal Sciences and Industry / Joel DeRouchey / Thirteen experiments using a total of 7,589 nursery and finishing pigs were conducted to evaluate the effects of deoxynivalenol (DON), feed additives and processing conditions on nursery pig growth performance. In addition, feed withdrawal and diet blending were evaluated in finishing pigs. Experiment 1 tested 3 feed additives in DON-contaminated diets with only Defusion Plus improving performance. Experiment 2 evaluated Biofix in both low- and high-DON diets and showed no effects on growth. Experiments 3 and 4 further evaluated levels of Defusion and the effects of pelleting and supplemental nutrients in DON-contaminated diets. Defusion improved growth in low-DON diets, but had variable effects in high DON diets. Pelleting DON-contaminated diets resulted in comparable growth to pigs fed positive control diets in meal form. In Exp. 5 and 6, pilot studies evaluated DON-detoxification using sodium metabisulfite (SMB) with hydrothermal treatment in both an autoclave and a pellet mill. These conditions reduced analyzed DON by as much as 89 and 75% for the autoclave and pellet mill, respectively. In Exp. 7 and 8, pelleting DON-contaminated diets with SMB improved growth. Experiments 9 and 10 evaluated feed-withdrawal time on carcass composition and economic returns. These experiments showed that pre-slaughter fasting for up to 36 h prior can be used to avoid weight discounts in heavyweight pigs without negatively impacting carcass composition and maintaining overall revenue. However, these advantages come with a potential reduction in carcass weight and increased incidence of leaking ingesta, which can result in condemned heads. Experiments 11, 12, and 13 compared phase-feeding to blending diets using an automated feed delivery system. These studies showed that corn-supplement blending is not economical and feeding diets blended to a Lys curve results in lower feed costs compared to phase-feeding, but due to reductions in growth and carcass weight, these savings do not translate into higher income over feed cost. Finally, Exp. 13 showed that over- and under-budgeting situations do not significantly influence overall returns, but pigs fed under-budgeted diets performed more closely to those fed correctly estimated feed budgets.

A presença de desoxinivalenol (DON) em produtos da moagem de trigo tem sido estudada, evidenciando que a contaminação ocorre em todas as frações. No Brasil, os limites máximos toleráveis (LMT) para DON em cereais e produtos de cereais são regulamentados pela Agência Nacional de Vigilância Sanitária - ANVISA. O entendimento da distribuição de DON nas frações da moagem do trigo é importante, pois pode fornecer base técnica para o gerenciamento desta contaminação e subsídios para as agências regulamentadoras no estabelecimento e ou revisão dos LMT. Este estudo teve como objetivo principal avaliar a distribuição de DON entre as frações obtidas da moagem experimental de grãos de trigo naturalmente contaminados. Trinta amostras de grãos de trigo (3-4 kg), naturalmente contaminadas com DON em concentrações que variaram de 350 a 4.150 ?g.kg-1, passaram por processo de limpeza (Labofix Brabender), seguido de condicionamento e moagem experimental (Brabender Quadrumat Senior). Quatro frações foram obtidas, de acordo com o tamanho das partículas: farelo (>=530 ?m), farelinho (>=195 e <=529 ?m), farinha de quebra (<=154 ?m) e farinha de redução (>=155 e <=194 ?m). As frações e o resíduo de limpeza foram avaliados quanto ao percentual obtido e concentração de DON. O DON foi extraído com H2O destilada, seguido de purificação em coluna de imunoafinidade e detecção/quantificação empregando cromatografia líquida de alta eficiência e detector de arranjo de diodos (UV-DAD). O percentual médio de resíduo de limpeza foi de 15,9% (6,9 a 23,2%). A redução média de DON nos grãos limpos foi 22,5% de (5,5 a 37,3%). A moagem experimental do trigo produziu em média 28,5% de farelo, 5,8% de farelinho, 27,9% de farinha de quebra, 37,8% de farinha de redução, totalizando 65,7% de farinha. As concentrações de DON verificadas no farelo e farelinho foram significativamente maiores quando comparadas com as farinhas (p<=0,05), as quais não diferiram entre si. A concentração relativa (CRel) de DON (relação entre a concentração da fração e do grão) indicou que a concentração de DON foi, em média, 73% maior no farelo e 35% maior no farelinho, comparada à concentração inicial nos grãos limpos. Na farinha de quebra, farinha de redução e farinha total a CRel foi em média 24%, 38% e 33% menor que nos grãos limpos, respectivamente. De acordo com os LMT para DON em farelo (2.000 ?g.kg-1) e farinha de trigo (1.750 ?g.kg-1), em vigência no Brasil, das 30 amostras avaliadas, 15 amostras de farelo (50%) e 1 amostra de farinha (3%) encontravam-se acima dos LMT. Considerando os LMT previstos pela legislação para 2017 (1.000 ?g.kg-1 para farelo e 750 ?g.kg-1 para farinha), 22 amostras de farelo (73%) e 16 amostras de farinha (53%) estariam acima dos LMT. Os resultados observados neste estudo demonstraram que a moagem experimental de grãos de trigo com contaminação igual a 3.000 ?g.kg-1, que será o LMT para grãos para posterior processamento em 2017, poderá acarretar a produção de farelos e farinhas com contaminação acima dos LMT previstos nesta mesma legislação. / The presence of deoxynivalenol (DON) in wheat milling products has been studied, showing that the contamination occurs in all streams. In Brazil, the maximum tolerable limits (MLT) for DON in cereals and cereal products are regulated by Agência Nacional de Vigilância Sanitária - ANVISA. The knowledge of DON distribution in wheat milling streams is important to provide the technical basis for the management of this contamination and could assist regulatory agencies to establish or review MTL. The main purpose of this study was to evaluate the fate of DON among experimental milling streams of naturally DON contaminated wheat kernels. Thirty wheat samples (3-4 kg), naturally DON contaminated (350-4.150 ?g.kg-1) were cleaned (Labofix Brabender), conditioned and milled (Brabender Senior Quadrumat mill). Four milled streams were obtained, according to the particle size: bran (>=530 ?m), shorts (>=195 e <=529 ?m), break flour (<=154 ?m) and reduction flour (>=155 e <=194 ?m). The streams and the screenings were evaluated for percentage and DON contents. DON analysis was performed with distilled H2O as extraction solvent, immunoaffinity column for cleanup and HPLC/diode array detector. The mean screening percentage obtained was 15,9% (6,9 to 23,2%). The mean reduction of DON in the cleaned wheat was 22,5% (5,5 to 37,3%). In the experimental milling of wheat it was obtained 28,5% of bran, 5,8% of shorts, 27,9% of break flour, and 37,8% of reduction flour, totaling 65,7% of flour. DON concentration obtained in bran and shorts were significantly higher when compared with the flours (p<=0,05), which did not differ from each other. The Relative Concentration (RConc) of DON (ratio between stream and cleaned wheat concentration) indicated that DON concentration was, on average, 73% higher in bran and 35% higher in shorts, compared to the initial concentration of the cleaned wheat, whereas in the break flour, reduction flour and total flour it was 24%, 38% and 33%, respectively, lower than the cleaned wheat. According to MTL for DON in bran (2.000 ?g.kg-1) and wheat flour (1.750 ?g.kg-1), in force in Brazil, the 30 samples evaluated, 15 samples of bran (50%) and 1 sample of flour (3%) were above the MLT. Considering the MTL provided in the legislation for 2017 (1.000 ?g.kg-1 for bran and 750 ?g.kg-1 for flour), 22 samples of bran (73%) and 16 samples of flour (53%) were above the MTL. The results of this study demonstrate that the experimental milling of wheat kernel contaminated at level of 3.000 ?g.kg-1 of DON, which will be the MTL for wheat kernels for further processing in 2017, may result in bran and flour contamination above the MTL provided in the same legislation.

O presente trabalho visou utilizar o perfil polifásico, envolvendo características fenotípicas e genotípicas, na identificação de Fusarium spp. isolados de grãos de trigo como também investigar a presença das principais toxinas: desoxinivalenol e zearalenona nos grãos de trigo de três regiões produtoras de trigo (São Paulo, Paraná e Rio Grande do Sul). Nossos resultados indicaram que os gêneros mais frequentes no trigo foram Alternaria, Fusarium e Epicoccum. A determinação do genótipo por qPCR demonstrou predomínio de 15-ADON seguido por NIV e 3-ADON. A quantificação de DNA demostrou que o perfil 15-ADON foi responsável por 96% de todo DNA quantificado, seguido por NIV com 3.84% e 3-ADON, com somente 0.06%, indicando que 15-ADON, é o principal genótipo de tricoteceno nos grãos de trigo nacional. A toxina desoxinivalenol foi detectada em todas amostras analisadas, com mediana de 323, 466 e 783 µg/kg em SP, PR e RS, respectivamente. A determinação de zearalenona demonstrou contaminação em 100%, 80% e 42% dos grãos dos Estados do RS, PR e SP e medianas de 843, 100 e 14 µg/kg, respectivamente. / The present work aimed to use a polyphasic approach, by phenotypic and genotypic characteristics for the identification of Fusarium strains from wheat grains as well to investigate the presence of deoxynivalenol and zearalenone on wheat grains from three wheat producers States (Parana, Rio Grande do Sul and Sao Paulo). Our results showed that the main genera found were Alternaria, Fusarium and Epicoccum. Genotype detection from qPCR revealed predominance of 15-ADON, followed by NIV and 3-ADON. The qPCR demonstrated that 15-ADON genotype was responsible for 96% of all DNA quantified, followed by NIV with 3.84% and 3-ADON with 0.06%, indicating that 15-ADON is the main trichothecene genotype from Brazilian wheat grains. The toxin deoxynivalenol was detected in all 150 samples analyzed from wheat grains, with medians of 323, 466 and 783 µg/kg in SP, PR and RS, respectively. The zearalenone determination showed contamination on 100%, 80% and 42% of wheat grains from RS, PR and SP State and medians of 843, 100 and 14 µg/kg, respectively.

Made available in DSpace on 2017-05-12T14:47:08Z (GMT). No. of bitstreams: 1 Tania_ Claudia Pintro.pdf: 1091369 bytes, checksum: f8c1d3904413167a06cca540a4fcaa7a (MD5) Previous issue date: 2015-02-12 / The objective of this study was to evaluate the effect of insect infestation Sitophilus zeamais on the physicochemical and microbiological quality in wheat grains, during storage in times of 0, 45, 90 and 135 days, in three temperatures: 20, 25 and 30 ºC. For the conduct of the experiment, wheat grain samples were collected (Triticum aestivum L.) of the cultivar BRS Tanager, produced in Western Paraná, from the harvest of 2013. For storage, the grains had water contents below 13%, being stored about 400 g in sealed glass containers with fabric type voil in order to facilitate gas exchange, totaling 72 containers stored in incubator BOD the temperature of 20, 25 and 30 ° C for 135 days. Of the total of containers, 36 were infested with 20 adults of Sitophilus zeamais and the other 36 were kept without infestation. Every 45 days, were samples for subsequent chemical-physical analysis (moisture, ash, and total protein population variation of insects), microbiological (filamentous fungi count and identification of fungi producing of deoxynivalenol) and quantifying of deoxynivalenol for the three temperatures at baseline and 135 days with and without infestation. The samples were evaluated by the use of analysis of variance (ANOVA) and for test of comparison of means the Tukey test was applied, with 5% level of significance. It was observed that, with increased storage time, increased population of insects, mainly at temperatures of 20 and 25° C, reducing the value of the weight of the hectolitre at both temperatures. In general, the water content was reduced with increasing temperature and time of the storage. The contents of protein, in the treatments with and without infestation, increased with storage time. In general, the ash content increased with time of storage in grain, with and without infestation in the three temperatures evaluated. The total count of CFU of filamentous fungi was greater for grains of wheat infested. The results obtained indicate that all samples of grain stored, analyzed in the initial time and 135 days, with and without infestation, for use at temperatures of 20, 25 and 30 &#7506;C, are within the maximum tolerable limit of 3000 &#956;g of deoxynivalenol per kg of wheat. / O objetivo deste trabalho foi avaliar o efeito da infestação de insetos Sitophilus zeamais sobre a qualidade físico-química e microbiológica de grãos de trigo, durante o armazenamento nos tempos de 0, 45, 90 e 135 dias e em três, temperaturas: 20, 25 e 30 ºC. Para a condução do experimento, foram coletadas amostras de grãos de trigo (Triticum aestivum L.) do cultivar BRS Tangará, produzido na região Oeste do Paraná e provenientes da safra de 2013. Para o armazenamento, os grãos apresentaram teores de água abaixo de 13%, sendo armazenados cerca de 400 g em recipientes de vidros, fechados com tecido tipo voil a fim de facilitar as trocas gasosas, totalizando 72 recipientes armazenados em câmara incubadora B.O.D. nas temperaturas de 20, 25 e 30 °C, no tempo de 135 dias. Do total de recipientes, 36 foram infestados com 20 insetos adultos de Sitophilus zeamais e os outros 36 foram mantidos sem infestação. A cada 45 dias, foram retiradas amostras, para posterior análise físico-química, (teor de água, cinzas, proteína total e variação populacional de insetos), microbiológicas (contagem de fungos filamentosos e identificação dos fungos produtores de desoxinivalenol) e de quantificação de desoxinivalenol, para as três temperaturas no tempo inicial e de 135 dias com e sem infestação. As amostras foram avaliadas pelo emprego da análise de variância (ANOVA) e para o teste de comparação de médias foi aplicado o teste de Tukey, com nível de 5% de significância. Observou-se que conforme aumentava o tempo de armazenamento aumentava também a população de insetos, principalmente para as temperaturas de 20 e 25 °C, reduzindo o valor do peso hectolitro em ambas as temperaturas. Em geral, o teor de água diminuiu quando ocorreu aumento de temperatura e tempo de armazenamento. Os teores de proteínas, nos tratamentos com e sem infestação, aumentaram com o tempo de armazenamento. Em geral, o teor de cinzas aumentou com o tempo de armazenamento nos grãos com e sem infestação, nas três temperaturas avaliadas. A contagem total de UFC de fungos filamentosos foi maior para grãos de trigo infestados. Os resultados obtidos mostram que todas as amostras de grãos armazenados, analisadas no tempo inicial e de 135 dias, com e sem infestação para as temperaturas de 20 25 e 30 &#7506;C, estão dentro do limite máximo tolerável de 3000 &#956;g de desoxinivalenol por kg de trigo

Dentre os desafios do cultivo de trigo, encontram-se as doenças fúngicas e a presença de micotoxinas. Estima-se que as perdas anuais nesse setor sejam bilionárias. Além dos prejuízos econômicos, há preocupações quanto a segurança alimentar humana e animal. Com o objetivo de avaliar a micobiota e a ocorrência de micotoxinas em diferentes estádios de maturação do trigo foram realizados estudos morfológicos, utilizando métodos morfológicos clássicos e moleculares, identificação do perfil genotípico dos fungos produtores de tricotecenos, por PCR em tempo real e determinação de deoxinivalenol (DON) e zearalenona (ZEA), utilizando a Cromatografia Líquida de Alta Eficiência acoplada à Espectrometria de Massas (LC-MS/MS). Os dados obtidos também foram correlacionados com os fatores climatológicos. Os resultados revelaram a predominância do gênero Fusarium nas amostras das duas safras, seguido de Alternaria e Epicoccum. A atividade de água média para os dois anos foram, 0,608 e 0,886, consecutivamente. Dentro do gênero Fusarium constatou-se predomínio do complexo de espécies Fusarium graminearum (CEFG), identificando F. graminearum sensu stricto, F. meridionale, F. cortaderiae e F. austroamericanum. Os genótipos dos isolados foram, na sua maioria, 15-ADON para F. graminearum s.s, seguido de NIV para as espécies F. meridionale, F. cortaderiae e F. austroamericanum e 3-ADON para as duas últimas. Por PCR em tempo real, a determinação do genótipo nos grãos de trigo nas safras 2014/15 e 2015/16 demonstraram predomínio de 100% e 93% para 15-ADON, respectivamente. A quantificação de DNA dos isolados do CEFG demostrou que o perfil 15-ADON foi responsável por mais de 95% nas duas safras, com valores irrisórios, menores que 3% para 3-ADON e NIV. Os níveis de contaminação pela micotoxina DON nas amostras foi de 100% nas duas safras, com valores variando entre 24,3 a 5530 &#956g/kg. Para ZEA os valores foram 30% e 96%, nas duas coletas consecutivas, variação entre 26,0 a 4360 &#956g/kg. Ambas as micotoxinas foram detectadas em concentrações acima do limite estabelecido pela legislação brasileira, 3000 &#956g/kg para DON e 400 &#956g/kg para ZEA. Os fatores climáticos obtiveram uma correlação positiva tanto na contaminação fúngica quanto no acúmulo de micotoxinas. Este estudo forneceu novas informações sobre a micobiota natural do trigo brasileiro, em diferentes etapas do cultivo, demonstrando a contaminação por DON e alta frequência de ZEA em amostras do provindas do Paraná. / The fungal diseases and the presence of mycotoxins in cereals cause annually, billionaire losses. In addition, there are concerns about human and animal food safety. Due to the wheat culture relevance in feed and foodstuff, the present work aimed to evaluate mycobiota and the occurrence of mycotoxins in different wheat maturation stages. In order to achieve these objectives, morphologic studies using classic and molecular methods, genotype profile and DNA quantification from trichothecene producers, by real-time PCR and determination of deoxynivalenol (DON) and zearalenone (ZEA), using High Performance Liquid Chromatography coupled to Mass Spectrometry (LC-MS/MS), were done. The obtained data were also correlated with the climatic factors. The results showed the predominance of the genus Fusarium followed by Alternaria and Epicoccum. The average water activity over two years were 0.608 and 0.886, consecutively. Within the genus Fusarium, it was identified a predominance of the Fusarium graminearum species complex (FGSC), identifying F. graminearum sensu stricto, F. meridionale, F. cortaderiae and F. austroamericanum. The genotypes of the isolates were 15-ADON for F. graminearum s.s, followed by NIV for the F. meridionale, F. cortaderiae and F. austroamericanum and 3-ADON for F. cortaderiae and F. austroamericanum. In real-time PCR, the determination of genotype in wheat grains in the 2014/15 and 2015/16 harvests showed a predominance of 100% and 93% for 15-ADON, respectively. The DNA quantification of the FGSC isolates showed that the 15-ADON profile was responsible for more than 95% in both harvests. The levels of DON mycotoxin contamination in the samples were 100%, with values ranging from 24.3 to 5530 &#956g/kg. For ZEA the values were 30% and 96%, in the two consecutive collections, ranging from 26,0 to 4360 &#956g/kg. Both mycotoxins were detected in concentrations above the limit established by Brazilian legislation, 3000 &#956g / kg for DON and 400 &#956g/kg for ZEA. The climatic factors obtained a positive correlation both in the fungal contamination and in the accumulation of mycotoxins. This study provided new information on the Brazilian native mycobiota to different stages of cultivation, demonstrating contamination by DON and high frequency of ZEA in samples from Paraná.