Occurrence of type A, B and D trichothecenes, zearalenone and stachybotrylactam in straw

An analytical protocol using high performance liquid chromatography (HPLC) was used to analyze samples of spring wheat, winter wheat, 2-row barley, and 6-row barley over a period of three years for the presence of five mycotoxins. These included deoxynivalenol, zearalenone, T-2 toxin, HT-2 toxin, and diacetoxyscirpenol. The protocol employed a single extraction step using acetonitrile-water and two cleanup procedures. One utilized a solid-phase extraction column and the other a charcoal-alumina column. Detection limits ranged from 0.02 to 0.15 micrograms of mycotoxin g-1 grain. Little T-2 toxin, HT-2 toxin, or diacetoxyscirpenol was found in the samples. Deoxynivalenol was detected in 53 to 62% of the samples tested and zearalenone in 25-29% of the samples. Several enzyme-linked immunosorbent assays were used for comparison purposes with deoxynivalenol, zearalenone, and T-2 toxin. These kits provided reliable qualitative, but not quantitative, data.

Samples of winter wheat (n =84), winter rye (46) and barley (29) were collected from the larger family farms and from partnerships in Lithuania just after the 1998 harvest. The number of samples collected from each region was proportional to the amount of grain produced in it. The levels of the Fusarium toxins deoxynivalenol (DON), 3-acetyl-DON, 15-acetyl-DON, nivalenol (NIV), fusarenon-X (4-acetyl-NIV), T-2 toxin, HT-2 toxin, 4,5-diacetoxyscirpenol (DAS), 1,5-monoacetoxyscirpenol (MAS) and scirpentriol in the grain were determined by gas chromatography with mass-selective detection (GC-MS). DON was most often detected in the wheat and rye samples and NIV in the barley samples. The concentrations found were lower than those causing acute or chronic toxic effects in livestock or humans. No fusarenon-X or 15-acetyl-DON was detected, and only small amounts of other trichothecenes were present. Climatic conditions in Lithuania in the summer of 1998 were slightly cooler and wetter than the average for the 1992-1996 but were close to the norm. Because the samples analysed were representative of grain produced for the market in seasons with normal weather, trichothecene contamination of grain from large family farms and partnerships would not be expected to be a problem in most years.

Animal feed can be contaminated with various mycotoxins. To ensure animal health and safe food and feed production, the European Commission has recommended increased monitoring of the co-occurrence of deoxynivalenol, zearalenone, ochratoxin A, fumonisin B1 and B2, T-2 and HT-2 toxin in feed. Thus, there is a need for an analytical method that enables their simultaneous detection and quantification. This paper describes the development and in-house validation of such a method, in which the mycotoxins were extracted from spiked and naturally contaminated cereal-based compound feed, corn and wheat. The extracts were divided into two aliquots where one was diluted and then analysed directly and the other was cleaned by using MultiSep®226 and then diluted and analysed. Separation and detection was achieved with LC-ESI-MS/MS by using a triple quadrupole instrument in the SRM mode. The precision (in terms of intra-day repeatability and inter-day reproducibility), accuracy, linearity, apparent recovery and expanded measurement uncertainty in feed, corn and wheat were evaluated. The LODs ranged from 1.0 to 72 μg/kg, and the LOQs ranged from 2.5 to 115 μg/kg. The apparent recovery was higher than 86% for all the mycotoxins, and the precision was better than that defined by the Horwitz equation for all concentrations. Proficiency test materials were analysed to assess the accuracy of the method, and the results were satisfactory for all seven mycotoxins. The method will be used to monitor the occurrence of these mycotoxins in products intended for animal feeding in Sweden.

Mycotoxins are toxic metabolites produced by certain types of fungi, causing pathological changes in humans and animals. The aim of this study was to assess the degree of contamination of selected cereal grains, bran and cereal products intended for children, with mycotoxins using GCxGC-TOF-MS technique. The study involved mycotoxins belonging to the type A and B trichothecenes group, including T-2 toxin (T-2), HT-2 toxin (HT-2), scirpenol (SCI), 15-monoacetoxyscirpenol (15-MAS), diacetoxyscirpenol (DAS), triacetoxyscirpenol (TAS), fusarenon-X (FUS-X), nivalenol (NIV), deoxynivalenol (DON), 3-acetyl-DON (3-Ac-DON), 15-acetyl-DON (15-Ac-DON). The study also assessed the effect of conditions in which the samples were stored, including temperature (6°C and 28°C) and time (14 and 28 days), on fungal growth and mycotoxin production. Among all studied compounds, only DAS and HT-2 toxins were detected in tested samples, with the exception of products intended for children. Measured HT-2 mycotoxin content in tested samples was in the range 83.9 - 196.4µg kg -1. Experiments with storage conditions showed a statistically significant increase in the HT-2 toxin level after 14 days of storage in all samples, irrespective of temperature. Prolonged storage (additional 14 days) did not cause significant changes in the HT-2 content. Further analyses showed a statistically significant effect of storage temperature on HT-2 toxin levels only in cereal products intended for children after both 14 and 28 days. Interestingly, lower temperature (6°C) was more optimal then higher temperature (28°C) for the HT-2 toxin production. No significant effect of storage temperature on HT-2 level was observed for cereal grains and bran.

Abstract. The aim of study was to detect the fungal and mycotoxins contamination of grain samples of oat, wheat and barley grown in four regions of Ural region (Kurgan, Sverdlovsk, Tyumen, Chelyabinsk) in 2017–2018. Methods. The infection of grain with fungi was analyzed using traditional mycological methods; the content of fungal DNA was determined by quantitative PCR; the presence and amounts of toxic secondary metabolites of fungi in the grain was detected by high performance liquid chromatography with mass spectrometry. Results. In the analyzed grain samples at least 10 species of Fusarium fungi were identified, among which F. sporotrichioides, F. avenaceum sensu lato and F. poae were found to be prevailing. The areas of Fusarium species that are atypical for the territory of Ural region were specified. F. graminearum was found in 14 % of the analyzed grain samples, and F. langsethiae was detected in three grain samples from the Sverdlovsk region. The DNA of F. poae was found in 48 % of grain samples, F. avenaceum DNA – in 39 %, F. sporotrichioides DNA – in 30 %, and F. graminearum DNA – in 29 % of analyzed grain samples. The content of mycotoxins in the grain samples ranged significantly depending on the crop and the geographical origin of the samples. One to seven mycotoxins were present in each contaminated grain sample. T-2 and HT-2 toxins were most common and were detected in 59 % of samples. Following to them beauvericin and deoxynivalenol were found in 34 % and 25 % of the grain samples, respectively. The excess of the maximum permissible level of T-2 toxin in 26 times was detected in grain of barley from the Chelyabinsk region Scientific novelty. For the first time, the information about the occurrence and the amounts of moniliformin and beauvericin, which are rarely analyzed in the grain, is provided. The significant connection between the content of DNA of dominant Fusarium species and the amount of the main mycotoxins produced by them in the grain were revealed.

Although the common oat (Avena sativa L.) is well known for its nutritional benefits, it carries the risk of contamination with mycotoxins due to its susceptibility to the growth of various fungi. The procurement of milling oats for food could become more difficult in the coming harvest years due to limited availability, specific quality requirements and the avoidance of mycotoxin contamination. In light of ongoing discussions in the European Commission on regulatory limits for certain mycotoxins including their modified forms, the purpose of this study was to improve the database on their occurrence in milling oats. In particular, we provide data on the predominantly occurring trichothecenes such as deoxynivalenol and its acetylated and modified derivatives (e.g. 3-acetyl-deoxynivalenol, 15-acetyl-deoxynivalenol and deoxynivalenol-3-glucoside) as well as on T-2 and HT-2 toxins. Additionally, the following mycotoxins were analysed: zearalenone, nivalenol, diacetoxyscirpenol, fusarenon-X, ochratoxin A, sterigmatocystin and aflatoxins B1, B2, G1 and G2. Oat samples, (n = 281) pre-selected for their physical properties and DON-content to be less than 1750 µg/kg from 11 European provenances, were analysed for 16 different mycotoxins by LC-MS/MS. Samples were collected from the years of harvest 2013 to 2019. High incidence rates above the limit of quantification of either 5 µg/kg for T-2 and HT-2 toxins or 10 µg/kg for deoxynivalenol were found (98.1, 94.7 and 91.4%, respectively). The mean concentration of the sum of T-2 and HT-2 toxins was 149 µg/kg. The highest level was found in an Irish sample containing 1290 µg/kg for the sum of T-2 and HT-2 toxins. The mean deoxynivalenol concentration was 289 µg/kg, while the highest level was 1414 µg/kg in a Swedish sample. Besides nivalenol other mycotoxins were only present in trace concentrations or not detected.

This study has examined the pattern of mycotoxin contamination of maize destined for animal feed in different global regions over a period of 3 years (2018–2020) with up to 1000+ samples analysed in each year. Overall, >75% of samples in each of the survey years were contaminated with multiple mycotoxins regardless of the global region (Europe, Africa, Asia, South Americas countries). Using LC-MS/MS, it was possible to quantify the relative contamination present in the samples in each year from the different regions of eight different mycotoxins including aflatoxin B1 (AFB1), ochratoxin A (OTA) deoxynivalenol (DON), fumonisin B1 (FB1) and B2, zearalenone (ZEA), T-2 and HT-2 toxins. The trends in mycotoxin contamination showed that there was a consistent contamination with DON in the 3 sampling years in all four regions. Interestingly, AFB1 contamination was prevalent in all regions in 2018, but more predominant in Europe and in 2019. In contrast, in 2020 it was found to be the major contaminant in Africa only. However, FB1 contamination of maize which was prevalent in Europe in 2018, became more prevalent in Asia and LATAM countries in 2019 and even in African maize in 2020. Comparisons of contamination with different mycotoxins in each of the years globally showed significant differences for AFB1, FB1, DON and ZEA between the different years. These results are discussed in relation to the trends of contamination of maize with mixtures of mycotoxins and the implication for their control in this key commodity used as an important ingredient in animal feed.

The amount of precipitation and its seasonal distribution affect the contamination of grain fodder with micromycetes. The use of grain from different crops as animal feed and in the food industry raises concerns about its quality and adherence to sanitary and medical standards. The aim of this study was to investigate the degree of micromycetes contamination in grain fodder (wheat, corn, barley) in the northwestern Black Sea region, taking into account weather conditions during 2017–2022. Methods. After harvesting, grain samples were taken and inoculated into Petri dishes containing potato dextrose agar to establish the total number of fungi and determine their species composition. The number of fungal spores in 1 g of feed was calculated using the method of serial dilutions. Micromycetes species were identified using classical methods based on cultural and morphological characteristics. Moisture conditions during the period of active vegetation, defined as having average daily temperatures above 10 °C, were assessed using the hydrothermal coefficient (HTC) of the Selyaninov method. The HTC was not calculated for months without active air temperatures above 10 °C. Results. In the northwestern Black Sea region of Ukraine, the moderate and dry period has increased from four to eight months, with an HTC of 0.4–0.8. The period of sufficient moisture (2018, 2021) with an HTC of 1.0–1.5 lasted five months. The amount of precipitation and its seasonal distribution in the northwestern Black Sea region affects the contamination of grain fodder with micromycetes. During the years of severe drought (2017, 2022), wheat grain was primarily contaminated with Aspergillus fungi, while corn and barley were contaminated with Penicillium. In the years of moderate drought (2019, 2020), grain feed was most contaminated with Penicillium fungi, and during the period of sufficient moisture (2018, 2021), it was contaminated with Penicillium and Fusarium fungi. Conclusions. The mycobiota of grain fodder in the northwestern Black Sea region of Ukraine is dependent on natural and climatic conditions. Filamentous fungi were the most commonly found in corn grain. During a period of severe drought, the grain was contaminated with fungi from the genera Penicillium (81%) and Aspergillus (78%). The other four fungal genera found were Fusarium (42%), Alternaria (31%), Mucor (16%), and Rhizopus (5%). In years with adequate moisture, the occurrence of Penicillium and Aspergillus fungi decreased by 17% and 14%, respectively, while the incidence of Fusarium fungi increased by 9%. During the period of sufficient moisture, wheat grain samples were primarily affected by fungi of the genera Penicillium (52%) and Aspergillus (45%). Fungi of the genera Rhizopus, Mucor, and Alternaria were isolated at a frequency ranging from 5% to 10%. In severe drought, barley and corn samples were primarily affected by fungi of the genera Penicillium (51%) and Aspergillus (42%). During the period of sufficient moisture, the frequency of isolation for Alternaria, Mucor, and Rhizopus fungi was 9%, 8%, and 4%, respectively.

Se estima que a nivel mundial se procesa setecientos mil millones de toneladas al año de grano de cacao, en contraste a su proceso productivo, solo se aprovecha las semillas, que representa un 20%-23% del fruto, la otra proporción del fruto como la mazorca o cáscara, placenta, cascarilla y jugo de mucílago son los principales subproductos y se pierde como desecho. Debido a la disponibilidad y su alto valor nutricional, se le está dando diferentes usos en nuevas áreas de aplicación. Por ello, esta revisión se centra en actualizar conocimientos sobre el uso de subproductos del cacao en la producción animal, nuevos enfoques su viabilidad y su sostenibilidad. En este contexto, los subproductos son utilizados en la alimentación en monogástricos y poligástricos, reportando resultados favorables y mayor rentabilidad, representando una alternativa viable y valiosa para la alimentación animal. Se ha comprobado que estos subproductos están compuestos por diversos compuestos bioactivos de interés farmacéutico, industrial y alimenticio, que ya le están dando nuevos usos y aplicaciones. Se anticipa que los subproductos de cacao serán ampliamente adoptados con otros fines. En consecuencia, se espera que en el futuro próximo su disponibilidad para la alimentación animal disminuya y sus precios aumenten.

Contamination of Fusarium moulds and subsequent production of mycotoxins is a common problem in cereals grown in Northern Europe. Oats is the third most important cereal crop in Sweden and is commonly contaminated by Fusarium. To date there are no published reports on the distribution of trichothecene-producing Fusaria in Swedish oats and correlation to T-2 and HT-2, the two most dominant toxins. Identification of Fusarium species by traditional methods requires specific skill and experience and quantification of specific species is not possible. Using modern molecular methods such as real-time PCR both identification and quantification of specific Fusarium species are possible directly from the food or feed samples. In this study, specific methods for Fusarium langsethiae, Fusarium sporotrichioides, Fusarium graminearum, Fusarium culmorum and Fusarium poae as well as for the total amount of DNA from trichothecene-producing species were evaluated with respect to precision, robustness and efficiency and then used in the analysis of oat samples. Sixty-two oat samples were taken from field trial harvests from 2006-2008 in the central and southern parts of Sweden and analysed for the presence of Fusarium species by using species-specific real-time PCR-based methods with TaqMan or SYBR Green biochemistry. The T-2 and HT-2 toxin content was analysed by gas chromatography with electron capture detection of the toxins' pentafluoropropyl derivatives. The results showed that F. langsethiae and F. poae were the most dominant Fusarium species among those tested in Swedish oats during this period. The DNA level of F. langsethiae correlated well with the level of T-2 and HT-2 toxin (r=0.7), indicating that this species was the most important producer of T-2 and HT-2 toxins in Swedish oats during the sampling period (2006 to 2008). It can also be concluded that real-time PCR is a powerful tool when studying Fusarium species distribution in oats.

Food security issues have become important as prices of staple grains have soared in the last few years. Climate change is expected to increase pressure on food supply/quality/sustainability worldwide. Changes in rainfall patterns, drought, temperature and CO 2 all impact on staple food production systems. It has been suggested that there will be “hotspots” in different regions of the world where the temperature may increase by +2–4 °C, where rainfall patterns and drought events may increase, resulting in more rapid desertification and significantly impacting on staple food crop yields. Thus, for staple cereals a doubling of CO 2 (350 to 700 μl/l) and an increase in temperature will have a penalty on yield and nutritional quality, while for rice this could increase crop biomass, although effects on yield and quality are less clear. Plant stress inevitably leads to increased susceptibility to fungal infection, pre- and post-harvest, and potential for increased contamination by mycotoxins (Magan et al., 2011; Wu et al., 2011). We have previously demonstrated that mycotoxigenic fungal activity can lead to small changes in dry matter losses in climate change type conditions, which would still exceed the legislative limits, certainly in Europe. It is thus important to consider the potential for increased contamination of cereals by Aspergillus flavus and aflatoxins and Fusarium verticillioides and fumonisins in maize; Fusarium species and trichothecenes A and B, such as T-2 and HT-2 toxins, zearelenone and deoxynivalenol in small grain cereals. We have harnessed our published ecological data on the marginal conditions for growth and mycotoxin production in relation to temperature and water availability for a range of Fusarium, Aspergillus and Penicillium species and tried to integrate this with other types of data including mould growth and molecular data to develop models which could be used to predict mycotoxin contamination and the relative risk in cereals under different climate change scenarios. Based on climate change scenarios there are expected to be hot spots of increased temperature and flooding in northern parts of India with southern parts being subjected to increased drought episodes with increased temperatures and irrigation becoming essential. This could have profound impacts on cereal production, yield and grain quality in northern parts of India. This could impact on food security and availability in some parts of the Indian sub-continent.

Soybean is considered an important source of human food and animal feed. Okara and tofu whey are the main by-products of soymilk and tofu production. The distribution of enzymes ?-amylase and lipoxygenase (Lox) from six soybean genotypes in protein extracts of okara and tofu as well as in soymilk and tofu whey was assessed. SDS-electrophoretic results showed that production process significantly affected high molecular mass protein fractions of soybean by-products. Low amounts of Lox in soymilk were registered, while in okara protein extracts and tofu whey this enzyme was present in trace. In tofu protein extracts, Lox was registered in higher quantity that might be the result of the aggregation process during formation of the tofu gel. Content of ?-amylase in all soy protein products was high and similar to the content in soybean genotype. Content of ?-amylase in okara depended on respective soybean genotype.

The scientific community has placed increasing importance on sustainability, leading to the exploration of alternative bedding materials to the widely used aspen wood chips. Bedding plays a critical role in ensuring the wellbeing of animals and the validity of experimental outcomes. Compared to the frequently evaluated biological materials, such as corncobs or spelts, synthetic materials such as plastic granules have been less investigated. We characterized two thermostable plastic materials as an alternative bedding material in C57BL/6J and BALB/c mice. We examined the impact of those bedding alternatives on physiological parameters, behavior, health status, and cage climate in static and digitally ventilated cages as well as the possibility of a reuse cycle after reprocessing. The mouse lines showed different locomotor activity, feeding, and nestbuilding behavior on plastic granules. While ammonia levels were constantly higher in static cages than in ventilated cages, blood parameters were unaltered, and health status was maintained during the reuse procedure. We show the use of plastic granulate as bedding material for mice that has the potential for processing and recycling. Further, we show that the material is accepted differently by the lines in the preference choice test without affecting their health or hygiene status.

The effect of sodium hydroxide treatment on the chemical composition, digestibility and digestible energy content of wheat, barley and oat straws

Metsulfuron methyl, its hydroxy metabolite (A1), and its glucose conjugate metabolite (A) were determined in several crops by liquid chromatography (LC) using a simple extraction and cleanup scheme. After the parent and 2 major metabolites were extracted from crops with methanol, metabolite A was enzymatically hydrolyzed to metabolite A1 with β-glucosidase. The treated extracts were cleaned up chromatographically, and then metsulfuron methyl and metabolite A1 were quantitatively determined by reversed-phase LC with UV detection at 254 nm. Recovery of metsulfuron methyl and its 2 metabolites was 90 ± 6% over a range of 0.005 to 0.4 ppm from fortified samples of brown grain, wheat, barley, sugarcane stalk, and oat straw. Method detection limits (MDLs) were 0.005 ppm for metsulfuron methyl, 0.006 ppm for metabolite A, and 0.003 ppm for metabolite A1 in such crops as brown grain, wheat, barley, and sugarcane, and 0.01 ppm for metsulfuron methyl, 0.015 ppm for metabolite A, and 0.01 ppm for metabolite A1 in oat straw. The MDLs were estimated on the basis of a signal-to-noise ratio of 5 within a confidence interval of 95%. The method has potential application for the analyses of these analytes in other crops and feeds and may be applicable to other sulfonylurea herbicides.