Nivalenol Research Articles

Exploratory real-time kinetic analysis of the cytotoxicity induced by maize silage mycotoxins in a calf intestinal epithelial cell line

In a temperate climate, the mycotoxins deoxynivalenol (DON), nivalenol (NIV), enniatin B (ENN B), mycophenolic acid (MPA), roquefortine C (ROC) and zearalenone (ZEN) are often found in maize silage. Although rumen microbiota are able to degrade some mycotoxins (e.g. DON), others are known to stay mainly intact (e.g. ROC). In addition, mycotoxin degradation can be hampered by a low ruminal pH or decrease in rumen microbial activity. Hence, these mycotoxins can reach the small intestine and exert a cytotoxic effect on intestinal epithelial cells. In this study, a real-time kinetic analysis of the cytotoxicity of these six mycotoxins and some of their metabolites (α- and β-zearalenol, α- and β-ZOL) was performed in a calf small intestinal epithelial cell line (CIEB). Confluency as well as the cell death parameters apoptosis and necrosis were determined to evaluate the mycotoxin-induced cytotoxicity. A combination of Annexin-V green and Cytotox red staining was used to determine early and late apoptosis as well as necrosis. Six different concentrations were tested ranging from 0.78 to 12.5 μM. Compared to cells not exposed to mycotoxins, DON and NIV exert a fast toxic effect with DON being more toxic than NIV within the first hours of incubation, whereas the inverse was observed at 16 h of incubation. On the other hand, MPA and ZEN induced increased Annexin V green positive cells within several hours of incubation with higher toxicity over time. Increased Annexin V green and Cytotox red positive cells were seen for ROC only at the highest concentration tested. For ENN B, increased Annexin V green positive cells were observed only after 12 h and α- and β-ZOL did not show cytotoxic effects. Hence, mycotoxin exposure causes either severe (DON and NIV) or more limited (ZEN, ROC, MPA, and ENN B) risk of bovine intestinal epithelial damage.

Detoxification and Excretion of Trichothecenes in Transgenic Arabidopsisthaliana Expressing Fusarium graminearum Trichothecene 3-O-acetyltransferase.

Fusarium graminearum, the causal agent of Fusarium head blight (FHB), produces trichothecenes including deoxynivalenol (DON), nivalenol (NIV), and 3,7,15-trihydroxy-12,13-epoxytrichothec-9-ene (NX-3). These toxins contaminate grains and cause profound health problems in humans and animals. To explore exploiting a fungal self-protection mechanism in plants, we examined the ability of F. graminearum trichothecene 3-O-acetyltransferase (FgTri101) to detoxify several key trichothecenes produced by F. graminearum: DON, 15-ADON, NX-3, and NIV. FgTri101 was cloned from F. graminearum and expressed in Arabidopsis plants. We compared the phytotoxic effects of purified DON, NIV, and NX-3 on the root growth of transgenic Arabidopsis expressing FgTri101. Compared to wild type and GUS controls, FgTri101 transgenic Arabidopsis plants displayed significantly longer root length on media containing DON and NX-3. Furthermore, we confirmed that the FgTri101 transgenic plants acetylated DON to 3-ADON, 15-ADON to 3,15-diADON, and NX-3 to NX-2, but did not acetylate NIV. Approximately 90% of the converted toxins were excreted into the media. Our study indicates that transgenic Arabidopsis expressing FgTri101 can provide plant protection by detoxifying trichothecenes and excreting the acetylated toxins out of plant cells. Characterization of plant transporters involved in trichothecene efflux will provide novel targets to reduce FHB and mycotoxin contamination in economically important plant crops.

Mycotoxin Occurrence and Risk Assessment in Gluten-Free Pasta through UHPLC-Q-Exactive Orbitrap MS.

Celiac disease (CD) is a genetic-based autoimmune disorder which is characterized by inflammation in the small intestinal mucosa due to the intolerance to gluten. Celiac people should consume products without gluten, which are elaborated mainly with maize or other cereals. Contamination of cereals with mycotoxins, such as fumonisins (FBs) and aflatoxins (AFs) is frequently reported worldwide. Therefore, food ingestion is the main source of mycotoxin exposure. A new analytical method was developed and validated for simultaneous analysis of 21 mycotoxins in gluten-free pasta, commonly consumed by celiac population as an alternative to conventional pasta. Ultrahigh-performance liquid chromatography coupled to quadrupole Orbitrap high-resolution mass spectrometry (UHPLC-Q-Exactive Orbitrap MS) was used for analyte separation and detection. The mycotoxins included in this work were those widely reported to occur in cereal samples, namely, ochratoxin-A (OTA), aflatoxins (AFB1, AFB2, AFG1 and AFG2), zearalenone (ZON), deoxynivalenol (DON), 3-acetyl-deoxynivalenol and 15-acetyl-deoxynivalenol (3-AcDON and 15-AcDON, respectively), nivalenol (NIV), neosolaniol (NEO), fusarenone-X, (FUS-X), T-2 toxin (T-2) and HT-2 toxin (HT-2), fumonisin B1 and B2 (FB1 and FB2, respectively), enniatins (ENN A, ENN A1, ENN B and ENN B1) and beauvericin (BEA). The validated method was successfully applied to 84 gluten-free pasta samples collected from several local markets of Campania region (Italy) during September to November 2020 to monitor the occurrence of mycotoxins and to assess the exposure to these food contaminants. A significant number of samples (95%) showed mycotoxin contamination, being Fusarium mycotoxins (FB1, ZON and DON) the most commonly detected ones. Regarding the risk assessment, the higher exposures were obtained for NIV, DON and FB1 for children and teenagers age group which can be explained due to their lower body weight.

The Effect of Foliar Fertilization with Micronutrients on Disease Severity and Mycotoxin Concentrations in the Grain of Winter Spelt (Triticum aestivum spp. spelta L.): A Case Study

The effect of mineral fertilization (NPK), foliar fertilization with micronutrients (Cu, Zn and Mn) and the NanoGro biostimulant on the severity of leaf, spike, stem base and root diseases in winter spelt cv. Schwabenkorn was evaluated in a field-plot experiment. A mycological analysis was performed and the content of Fusarium mycotoxins in grain was determined. Mineral fertilization (NPK), foliar fertilization with micronutrients and the NanoGro biostimulant exerted varied effects on the severity of Septoria leaf blotch and Septoria glume blotch, they promoted the spread of brown rust (excluding the NPK + NanoGro treatment) and inhibited the spread of black head mold (excluding the NPK treatment), eyespot and Fusarium foot and root rot (excluding the NPK + Mn treatment). Fertilization had no influence on grain yield or the content of Fusarium mycotoxins in grain. The concentrations of deoxynivalenol (DON), nivalenol (NIV), zearalenone (ZEA) and fumonisins FB1 + FB2 did not exceed the maximum permissible levels (except for DON in NPK + Cu + Zn + Mn and NPK + NanoGro treatments). Throughout the experiment, ergosterol (ERG) concentrations were highest in the grain of unfertilized spelt plants.

Fusarium cerealis causing Fusarium head blight of durum wheat and its associated mycotoxins

Fusarium Head Blight (FHB) is a very important fungal disease that affects small grain cereals worldwide. This disease not only causes yield loses but also crops contamination with mycotoxins such as deoxynivalenol (DON) and nivalenol (NIV). Species within the Fusarium graminearum species complex have been described as the main causal agents of this disease, however lately there have been few reports of Fusarium cerealis causing the disease in wheat and barley in different parts of the world. This study evaluated the aggressiveness of F. cerealis to durum wheat cultivars and also mycotoxin production in planta. Moreover, the mycotoxin profile of F. cerealis strains was characterized molecularly and chemically. All durum wheat cultivars showed typical FHB symptoms but the disease severity varied among them in levels up to 66%. In addition, seventeen different compounds were detected in the infected heads including DON, NIV and nivalenol-3-β-d-glucose (NIV3G). NIV was detected in all cultivars and was the most produced mycotoxin with levels ranging from 1.04 to 6.8 mg/kg. On the other hand, the molecular analysis of F. cerealis strains showed that all of them possessed NIV genotype while the chemical assessment showed that the strains were able to produce not only this toxin in vitro but also DON, zearalenone and other twenty-one secondary metabolites. The increasing incidence of F. cerealis and the possible contamination of crops with the mycotoxins that it produces are of great concern for food security and world cereal trade since it has been reported that NIV is more toxic for humans and animals than DON.

Occurrence of major type-B trichothecenes and deoxynivalenol-3-glucoside in cereal-based products from Korea

A total of 67 cereal samples consisting of conventionally and organically produced corn, wheat and rice were analyzed by ultra-performance liquid chromatography-tandem mass spectrometry to monitor the occurrence of type-B trichothecenes (deoxynivalenol (DON), nivalenol (NIV), 3-acetyldeoxynivalenol (3ADON), 15-acetyldeoxynivalenol (15ADON), fusarenon-X (FUS-X)), and deoxynivalenol-3-glucoside (D3G). The incidence rates were 94%, 88%, 86.5%, 31.3%, 29.8%, and 61.2% for DON, D3G, NIV, 3ADON, 15ADON, and FUS-X, respectively. The highest contamination levels of DON, D3G, NIV, 3ADON, and 15ADON were found in corn, followed by wheat and rice, whereas FUS-X exhibited high contamination in rice. Moreover, significant differences were observed between conventionally and organically produced corn samples, which revealed higher incidence and contamination levels of the target mycotoxins in organic samples rather than in conventional samples. A total of 93.6% of all DON-contaminated samples showed the transformation of DON to its glucosylated form, D3G, with average conversion rates of 14%, 17%, and 28% in rice, wheat, and corn samples, respectively. In addition, the co-occurrence of two or more different mycotoxins was observed in a high percentage of the analyzed samples. Therefore, the results indicated that a vigilant attitude should be adopted to reduce human exposure to all of these toxic compounds.

Determination of Zearalenone and Trichothecenes, Including Deoxynivalenol and Its Acetylated Derivatives, Nivalenol, T-2 and HT-2 Toxins, in Wheat and Wheat Products by LC-MS/MS: A Collaborative Study.

An analytical method for the simultaneous determination of trichothecenes—namely, nivalenol (NIV), deoxynivalenol (DON) and its acetylated derivatives (3- and 15-acetyl-DON), T-2 and HT-2 toxins—and zearalenone (ZEN) in wheat, wheat flour, and wheat crackers was validated through a collaborative study involving 15 participants from 10 countries. The validation study, performed within the M/520 standardization mandate of the European Commission, was carried out according to the IUPAC (International Union of Pure and Applied Chemistry) International Harmonized Protocol. The method was based on mycotoxin extraction from the homogenized sample material with a mixture of acetonitrile-water followed by purification and concentration on a solid phase extraction column. High-performance liquid chromatography coupled with tandem mass spectrometry was used for mycotoxin detection, using isotopically labelled mycotoxins as internal standards. The tested contamination ranges were from 27.7 to 378 μg/kg for NIV, from 234 to 2420 μg/kg for DON, from 18.5 to 137 μg/kg for 3-acetyl-DON, from 11.4 to 142 μg/kg for 15-acetyl-DON, from 2.1 to 37.6 μg/kg for T-2 toxin, from 6.6 to 134 μg/kg for HT-2 toxin, and from 31.6 to 230 μg/kg for ZEN. Recoveries were in the range 71–97% with the lowest values for NIV, the most polar mycotoxin. The relative standard deviation for repeatability (RSDr) was in the range of 2.2–34%, while the relative standard deviation for reproducibility (RSDR) was between 6.4% and 45%. The HorRat values ranged from 0.4 to 2.0. The results of the collaborative study showed that the candidate method is fit for the purpose of enforcing the legislative limits of the major Fusarium toxins in wheat and wheat-based products.

A Comparative Transcriptome Analysis, Conserved Regulatory Elements and Associated Transcription Factors Related to Accumulation of Fusariotoxins in Grain of Rye (Secale cereale L.) Hybrids.

Detoxification of fusariotoxin is a type V Fusarium head blight (FHB) resistance and is considered a component of type II resistance, which is related to the spread of infection within spikes. Understanding this type of resistance is vital for FHB resistance, but to date, nothing is known about candidate genes that confer this resistance in rye due to scarce genomic resources. In this study, we generated a transcriptomic resource. The molecular response was mined through a comprehensive transcriptomic analysis of two rye hybrids differing in the build-up of fusariotoxin contents in grain upon pathogen infection. Gene mining identified candidate genes and pathways contributing to the detoxification of fusariotoxins in rye. Moreover, we found cis regulatory elements in the promoters of identified genes and linked them to transcription factors. In the fusariotoxin analysis, we found that grain from the Nordic seed rye hybrid “Helltop” accumulated 4 times higher concentrations of deoxynivalenol (DON), 9 times higher nivalenol (NIV), and 28 times higher of zearalenone (ZEN) than that of the hybrid “DH372” after artificial inoculation under field conditions. In the transcriptome analysis, we identified 6675 and 5151 differentially expressed genes (DEGs) in DH372 and Helltop, respectively, compared to non-inoculated control plants. A Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that DEGs were associated with glycolysis and the mechanistic target of rapamycin (mTOR) signaling pathway in Helltop, whereas carbon fixation in photosynthesis organisms were represented in DH372. The gene ontology (GO) enrichment and gene set enrichment analysis (GSEA) of DEGs lead to identification of the metabolic and biosynthetic processes of peptides and amides in DH372, whereas photosynthesis, negative regulation of catalytic activity, and protein-chromophore linkage were the significant pathways in Helltop. In the process of gene mining, we found four genes that were known to be involved in FHB resistance in wheat and that were differentially expressed after infection only in DH372 but not in Helltop. Based on our results, we assume that DH372 employed a specific response to pathogen infection that led to detoxification of fusariotoxin and prevented their accumulation in grain. Our results indicate that DH372 might resist the accumulation of fusariotoxin through activation of the glycolysis and drug metabolism via cytochrome P450. The identified genes in DH372 might be regulated by the WRKY family transcription factors as associated cis regulatory elements found in the in silico analysis. The results of this study will help rye breeders to develop strategies against type V FHB.

Mycotoxins produced by fusarium species associated with maize ear rot in Iran

Mycotoxins contamination is one of the most important problems worldwide in maize that can cause serious threat for human and animal health. The aim of this study was to determine the ability of Fusarium species associated with maize ear rot to produce diverse mycotoxins. The results showed, One out of three isolates of F. subglutinans produced detectable level of Beauvericin (BEA); the only isolate of F. temperatum produced 302 µg/g of BEA. Two out of five isolates of F. redolens produced enniatin B1and four isolates of this species produced high levels of BEA. As well, 21 isolates of Fusarium incarnatum-equiseti species complex (FIESC) and one isolates of F. brachygibbosum were evaluated for production of trichothecens (T-2 toxin, HT-2 toxin, Diacetoxyscirpenol (DAS), nivalenol (NIV) and deoxynivalenol (DON)), zearalenone (ZEN), enniatins (ENN A, A1, B and B1) and BEA by HPLC. Production of trichothecens, ZEN and BEA with one, three and four isolates of FIESC, respectively were observed and one isolate of F. brachygibbosum only produced detectable level of DAS and ENN B1. Analyzing of moniliformin production of F. proliferatum (26 isolates), FIESC (21 isolates) and F.thapsinum (10 isolates) showed none of them produce this toxin. These results revealed the ability of Fusarium spp. from maize to produce a varied range of mycotoxins which are harmful to human and animal’s health. Therefore, the occurrence of such broad number of different species on Iranian maize could be reason of great concern because of the toxigenic risk associated to these species.

Mitigation of nivalenol using alcoholic fermentation and magnetic field application

This study aimed at evaluating mitigation of nivalenol (NIV) in alcoholic fermentation with magnetic field application (MF). Mitigation was related to both the glutathione (GSH) redox molecule and the enzyme peroxidase (PO), which were synthesized by Saccharomyces cerevisiae US-05. Conditions under evaluation were NIV (0.2 µg mL−1), MF application (35 mT) and simultaneous use of mycotoxin and MF. The GSH content and the PO activity were increased when the culture contained NIV and the alcohol profile was altered after 48 h of fermentation. At the end of the alcoholic fermentation, NIV was mitigated by 56.5%. Therefore, this process is a promising method to reduce contamination by NIV, although the mycotoxin affects the chemical characteristics of the final product.

Development of an Improved Method of Sample Extraction and Quantitation of Multi-Mycotoxin in Feed by LC-MS/MS.

A multi-mycotoxin chromatographic method was developed and validated for the simultaneous quantitation of aflatoxins (AFB1, AFB2, AFG1 and AFG2), ochratoxin A (OTA), zearalenone (ZON), deoxynivalenol (DON), nivalenol (NIV), diacetoxyscirpenol (DAS), fumonisins (FB1, FB2 and FB3), T-2 toxin (T-2) and HT-2 toxin (HT-2) in feed. The three most popular sample preparation techniques for determination of mycotoxins have been evaluated, and the method with highest recoveries was selected and optimized. This modified QuEChERS (quick, easy, cheap, effective, rugged and safe) approach was based on the extraction with acetonitrile, salting-out and cleanup with lipid removal. A reconstitution process in methanol/water was used to improve the MS responses and then the extracts were analyzed by LC-MS/MS. In this method, the recovery range is 70–100% for DON, DAS, FB1, FB2, FB3, HT-2, T-2, OTA, ZON, AFG1, AFG2, AFB1 and AFB2 and 55% for NIV in the spike range of 2–80 µg/kg. Method robustness was determined with acceptable z-scores in proficiency tests and validation experiments.

Cytotoxic effects of individual and combined sterigmatocystin and nivalenol on liver hepatocellular carcinoma cells

Since humans are exposed to different mycotoxins through daily intake, there is increasing concern about the adverse effects of the interactions between them. Cytotoxicity of sterigmatocystin (STE) and nivalenol (NIV) alone and in combination in human hepatocarcinoma (HepG2) cells was evaluated by MTT assay. Furthermore, ROS production and alteration of ΔΨm as mechanisms of action were assessed. Cells were treated with concentrations ranging from 0.15 to 5 μM for NIV and from 0.78 to 50 μM for STE individually and in binary combinations. The combination ratio between the mixture STE + NIV was 10:1. The IC50 values of NIV ranged from 0.96 to 0.66 μM, whereas no IC50 values were obtained for STE at any time tested. For the combinations studied, synergistic, antagonistic and addictive effects were obtained with the two type of analyses performed, the isobologram analysis and the Combenefit method. No relevant effects on ROS and ΔΨm were observed. In conclusion, predictive models based on combination data could help to better understand the interactions between mycotoxins and their implications in food safety assessment. However, a further analysis of the molecular mechanism underlying these interactive effects is required.

Evaluation of the Efficacy of Mycotoxin Modifiers and Mycotoxin Binders by Using an In Vitro Rumen Model as a First Screening Tool.

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 α-zearalenol (α-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.

Analysis of Fusarium graminearum Species Complex from Freshly Harvested Rice in Jiangsu Province (China).

Members of Fusarium graminearum species complex (FGSC) are the major pathogens that cause Fusarium head blight (FHB) in cereals worldwide. Symptoms of FHB on rice, including dark staining or browning of rice glumes, were recently observed in Jiangsu Province, China. To improve our understanding of the pathogens involved, 201 FGSC isolates were obtained from freshly harvested rice samples and identified by phylogenetic analyses. Among the 201 FGSC isolates, 196 were F. asiaticum and the remaining 5 were F. graminearum. Trichothecene chemotype and chemical analyses showed that 68.4% of the F. asiaticum isolates were the 3-acetyldeoxynivalenol (3ADON) chemotype and the remainder were the nivalenol (NIV) chemotype. All of the F. graminearum isolates were the 15-acetyldeoxynivalenol chemotype. Pathogenicity assays showed that both the 3ADON and NIV chemotypes of F. asiaticum could infect wheat and rice spikes. FHB severity and trichothecene toxin analysis revealed that F. asiaticum with the NIV chemotype was less aggressive than that with the 3ADON chemotype in wheat, while the NIV-producing strains were more virulent than the 3ADON-producing strains in rice. F. asiaticum isolates with different chemotypes did not show significant differences in mycelial growth, sporulation, conidial dimensions, or perithecial production. These findings would provide useful information for developing management strategies for the control of FHB in China.

Characterization of the Fusarium sambucinum species complex and detection of multiple mycotoxins in Brazilian barley samples

This study investigated the fungal diversity in Brazilian barley samples, focusing on the Fusarium sambucinum species complex and the presence of multiple mycotoxins: aflatoxins B1, B2, G1, G2 beauvericin (BEA), enniatins (ENNs) A, A1, B, and B1, deoxynivalenol (DON), fumonisins (FB) B1 and B2, HT-2 and T-2 toxins, nivalenol (NIV) and ochratoxin A (OTA) from two different regions, São Paulo (SP) and Rio Grande do Sul (RS). The majority of the isolates belonged to the Fusarium sambucinum species complex (FSAMSC), with F. graminearum s.s. characterized as the major contaminant. F. meridionale and F. poae were the second most frequent fungi isolated from SP and RS, respectively. All of the F. graminearum s.s. isolates demonstrated 15-ADON genotype, whereas F. poae and F. meridionale were all NIV. The majority of the F. cortaderiae isolates were NIV, with only one 3-ADON genotype. Mycotoxin analysis revealed that none of the samples were contaminated by aflatoxins, OTA, FB2 and type A trichothecenes, however, all of the samples were contaminated with at least one Fusarium toxin. Contamination by DON, ZEA, ENNB and ENNB1 levels were significantly higher in RS. Co-contamination of BEA, DON, ENNs, NIV and ZEA in 18.5% and 24.2% of the analyzed samples was observed, from SP and RS respectively. More than 20% of the samples from RS presented DON and ZEA levels above the regulations established by Europe and Brazil. The results provide further information on the FSAMSC from South America and detected multiple Fusarium toxins in barley samples. This highlights the importance for further studies on the possible interactions of these mycotoxins in order to determine potential risks to animal health.

First Report of Fusarium Head Blight Caused by Fusarium meridionale in Rice in China

First Report of Fusarium Head Blight Caused by <i>Fusarium meridionale</i> in Rice in China

Population Genomic Analysis Reveals a Highly Conserved Mitochondrial Genome in Fusarium asiaticum.

Fusarium asiaticum is one of the pivotal members of the Fusarium graminearum species complex (FGSC) causing Fusarium head blight (FHB) on wheat, barley and rice in large parts of Asia. Besides resulting in yield losses, FHB also causes the accumulation of mycotoxins such as nivalenol (NIV) and deoxynivalenol (DON). The aim of this study was to conduct population studies on F. asiaticum from Southern China through mitochondrial genome analyses. All strains were isolated from wheat or rice from several geographic areas in seven provinces in Southern China. Based on geographic location and host, 210 isolates were selected for next generation sequencing, and their mitogenomes were assembled by GRAbB and annotated to explore the mitochondrial genome variability of F. asiaticum. The F. asiaticum mitogenome proves extremely conserved and variation is mainly caused by absence/presence of introns harboring homing endonuclease genes. These variations could be utilized to develop molecular markers for track and trace of migrations within and between populations. This study illustrates how mitochondrial introns can be used as markers for population genetic analysis. SNP analysis demonstrate the occurrence of mitochondrial recombination in F. asiaticum as was previously found for F. oxysporum and implied for F. graminearum. Furthermore, varying degrees of genetic diversity and recombination showed a high association with different geographic regions as well as with cropping systems. The mitogenome of F. graminearum showed a much higher SNP diversity while the interspecies intron variation showed no evidence of gene flow between the two closely related and sexual compatible species.

Multiple Fungal Metabolites Including Mycotoxins in Naturally Infected and Fusarium-Inoculated Wheat Samples.

In this study, the occurrence of multiple fungal metabolites including mycotoxins was determined in four different winter wheat varieties in a field experiment in Croatia. One group was naturally infected, while the second group was inoculated with a Fusarium graminearum and F. culmorum mixture to simulate a worst-case infection scenario. Data on the multiple fungal metabolites including mycotoxins were acquired with liquid chromatography with mass spectrometry (LC-MS/MS) multi-(myco)toxin method. In total, 36 different fungal metabolites were quantified in this study: the Fusarium mycotoxins deoxynivalenol (DON), DON-3-glucoside (D3G), 3-acetyldeoxynivalenol (3-ADON), culmorin (CULM), 15-hydroxyculmorin, 5-hydroxyculmorin, aurofusarin, rubrofusarin, enniatin (Enn) A, Enn A1, Enn B, Enn B1, Enn B2, Enn B3, fumonisin B1, fumonisin B2, chrysogin, zearalenone (ZEN), moniliformin (MON), nivalenol (NIV), siccanol, equisetin, beauvericin (BEA), and antibiotic Y; the Alternaria mycotoxins alternariol, alternariolmethylether, altersetin, infectopyron, tentoxin, tenuazonic acid; the Aspergillus mycotoxin kojic acid; unspecific metabolites butenolid, brevianamid F, cyclo(L-Pro-L-Tyr), cyclo(L-Pro-L-Val), and tryptophol. The most abundant mycotoxins in the inoculated and naturally contaminated samples, respectively, were found to occur at the following average concentrations: DON (19,122/1504 µg/kg), CULM (6109/1010 µg/kg), 15-hydroxyculmorin (56,022/1301 µg/kg), 5-hydroxyculmorin (21,219/863 µg/kg), aurofusarin (43,496/1266 µg/kg). Compared to naturally-infected samples, Fusarium inoculations at the flowering stage increased the concentrations of all Fusarium mycotoxins, except enniatins and siccanol in Ficko, the Aspergillus metabolite kojic acid, the Alternaria mycotoxin altersetin, and unspecific metabolites brevianamid F, butenolid, cyclo(L-Pro-L-Tyr), and cyclo(L-Pro-L-Val). In contrast to these findings, because of possible antagonistic actions, Fusarium inoculation decreased the concentrations of the Alternaria toxins alternariol, alternariolmethylether, infectopyron, tentoxin, tenuazonic acid, as well as the concentration of the nonspecific metabolite tryptophol.

Application of Artificial Neural Networks to Analyze the Concentration of Ferulic Acid, Deoxynivalenol, and Nivalenol in Winter Wheat Grain

Biotic stress, which includes infection by pathogenic fungi, causes losses of wheat yield in terms of quantity and quality. Ear Fusarium is caused by strains of F. graminearum and F. culmorum, which can produce mycotoxins—deoxynivalenol (DON) and nivalenol (NIV). One of the wheat’s defense mechanisms against stressors is the activation of biosynthesis pathways of antioxidant compounds, including ferulic acid. The aim of the study was to conduct pilot studies on the basis of which neural models were created that would examine the impact of the variety and weather conditions on the concentration of ferulic acid, and link its content with the concentration of deoxynivalenol and nivalenol. The plant material was 23 winter wheat genotypes with different Fusarium resistance. The field experiment was conducted in 2011–2013 in Poland in three experimental combinations, namely: with full chemical protection; without chemical protection, but infested with natural disease (control); and in the absence of fungicidal protection, with artificial inoculation by genus Fusarium fungi. As a result of the pilot studies, three neural models—FERUANN analytical models (ferulic acid content), DONANN (deoxynivalenol content) and NIVANN (nivalenol content)—were produced. Each model was based on 14 independent features, 12 of which were in the form of quantitative data, and the other two were presented as qualitative data. The structure of the created models was based on an artificial neural network (ANN) of the multilayer perceptron (MLP) with two hidden layers. The sensitivity analysis of the neural network showed the two most important features determining the concentration of ferulic acid, deoxynivalenol, and nivalenol in winter wheat seeds. These are the experiment variant (VAR) and winter wheat variety (VOW).

Regional and field-specific differences in Fusarium species and mycotoxins associated with blighted North Carolina wheat

Worldwide, while Fusarium graminearum is the main causal species of Fusarium head blight (FHB) in small-grain cereals, a diversity of FHB-causing species belonging to different species complexes has been found in most countries. In the U.S., FHB surveys have focused on the Fusarium graminearum species complex (FGSC) and the frequencies of 3-ADON, 15-ADON, and nivalenol (NIV) chemotypes. A large-scale survey was undertaken across the state of North Carolina in 2014 to explore the frequency and distribution of F. graminearum capable of producing NIV, which is not monitored at grain intake points. Symptomatic wheat spikes were sampled from 59 wheat fields in 24 counties located in three agronomic zones typical of several states east of the Appalachian Mountains: Piedmont, Coastal Plain, and Tidewater. Altogether, 2197 isolates were identified to species using DNA sequence-based methods. Surprisingly, although F. graminearum was the majority species detected, species in the Fusarium tricinctum species complex (FTSC) that produce “emerging mycotoxins” were frequent, and even dominant in some fields. The FTSC percentage was 50–100% in four fields, 30–49% in five fields, 20–29% in five fields, and < 20% in the remaining 45 fields. FTSC species were at significantly higher frequency in the Coastal Plain than in the Piedmont or Tidewater (P < .05). Moniliformin concentrations in samples ranged from 0.0 to 38.7 μg g−1. NIV producing isolates were rare statewide (2.2%), and never >12% in a single field, indicating that routine testing for NIV is probably unnecessary. The patchy distribution of FTSC species in wheat crops demonstrated the need to investigate the potential importance of their mycotoxins and the factors that allow them to sometimes outcompete trichothecene producers. An increased sampling intensity of wheat fields led to the unexpected discovery of a minority FHB-causing population.