Overproduction of mycotoxin biosynthetic enzymes triggers Fusarium toxisome-shaped structure formation via endoplasmic reticulum remodeling.

Mycotoxin deoxynivalenol (DON) produced by the Fusarium graminearum complex is highly toxic to animal and human health. During DON synthesis, the endoplasmic reticulum (ER) of F. graminearum is intensively reorganized, from thin reticular structure to thickened spherical and crescent structure, which was referred to as "DON toxisome". However, the underlying mechanism of how the ER is reorganized into toxisome remains unknown. In this study, we discovered that overproduction of ER-localized DON biosynthetic enzyme Tri4 or Tri1, or intrinsic ER-resident membrane proteins FgHmr1 and FgCnx was sufficient to induce toxisome-shaped structure (TSS) formation under non-toxin-inducing conditions. Moreover, heterologous overexpression of Tri1 and Tri4 proteins in non-DON-producing fungi F. oxysporum f. sp. lycopersici and F. fujikuroi also led to TSS formation. In addition, we found that the high osmolarity glycerol (HOG), but not the unfolded protein response (UPR) signaling pathway was involved in the assembly of ER into TSS. By using toxisome as a biomarker, we screened and identified a novel chemical which exhibited high inhibitory activity against toxisome formation and DON biosynthesis, and inhibited Fusarium growth species-specifically. Taken together, this study demonstrated that the essence of ER remodeling into toxisome structure is a response to the overproduction of ER-localized DON biosynthetic enzymes, providing a novel pathway for management of mycotoxin contamination.

Role of ferroptosis in food-borne mycotoxin-induced toxicities.

Contamination by toxic substances is a major global food safety issue, which poses a serious threat to human health. Mycotoxins are major class of food contaminants, mainly including aflatoxins (AFs), zearalenone (ZON), deoxynivalenol (DON), ochratoxin A (OTA), fumonisins (FBs) and patulin (PAT). Ferroptosis is a newly identified iron-dependent form of programmed or regulated cell death, which has been found to be involved in diverse pathological conditions. Recently, a growing body of evidence has shown that ferroptosis is implicated in the toxicities induced by certain types of food-borne mycotoxins, which provides novel mechanistic insights into mycotoxin-induced toxicities and paves the way for developing ferroptosis-based strategy to combat against toxicities of mycotoxins. In this review article, we summarize the key findings on the involvement of ferroptosis in mycotoxin-induced toxicities and propose issues that need to be addressed in future studies for better utilization of ferroptosis-based approach to manage the toxic effects of mycotoxin contamination.

Photonic Microbead Array Digital Time-Resolved Fluorescence Ultrasensitive Platform for Simultaneous Detection of Multiple Mycotoxins.

Limitations in the sensitivity, linear detection range, and cross-reaction of lateral flow immunoassays mainly hamper their application in rapid screening for multiple targets. In this work, we designed a new time-resolved fluorescence immunoassay (TRFIA) platform to overcome these limitations. This platform uses europium chelate polystyrene (PS@Eu) nanoparticles conjugated with monoclonal antibodies to sense multiple mycotoxins. We employed a competitive TRFIA protocol in which the conjugated PS@Eu was used on the surfaces of photonic microbead arrays (PMAs). The TRFIA signal of PMAs on the pad was recorded with the digital time-resolved fluorescence reader. The developed TRFIA shows wide detection linear ranges (0.01-1000 ng/mL for DON, 0.1-100 ng/mL for OTA, and 0.01-100 ng/mL for AFB1), low limits of detection (LODs) (7.9 pg/mL for DON, 18 pg/mL for OTA, and 7.7 pg/mL for AFB1), good specificity, good recovery ratios (76.68-117.26%), and good reproducibility in grain samples. The simulated fluorescence enhancement effect of PMA indicated that the electric field distribution on the surface of PS@Eu on PMA is twice higher than that on the surface of PS@Eu. The new TRFIA for three kinds of mycotoxins was 1000-fold more sensitive than the classical TRFIA, and it has great potential application in rapid screening for multiple targets.

Genetic mapping of deoxynivalenol and fusarium damaged kernel resistance in an adapted durum wheat population.

BACKGROUND: Fusarium head blight (FHB) infection results in Fusarium damaged kernels (FDK) and deoxynivalenol (DON) contamination that are downgrading factors at the Canadian elevators. Durum wheat (Triticum turgidum L. var. durum Desf.) is particularly susceptible to FHB and most of the adapted Canadian durum wheat cultivars are susceptible to moderately susceptible to this disease. However, the durum line DT696 is less susceptible to FHB than commercially grown cultivars. Little is known about genetic variation for durum wheat ability to resist FDK infection and DON accumulation. This study was undertaken to map genetic loci conferring resistance to DON and FDK resistance using a SNP high-density genetic map of a DT707/DT696 DH population and to identify SNP markers useful in marker-assisted breeding. One hundred twenty lines were grown in corn spawn inoculated nurseries near Morden, MB in 2015, 2016 and 2017 and the harvested seeds were evaluated for DON. The genetic map of the population was used in quantitative trait locus analysis performed with MapQTL.6® software. RESULTS: Four DON accumulation resistance QTL detected in two of the three years were identified on chromosomes 1 A, 5 A (2 loci) and 7 A and two FDK resistance QTL were identified on chromosomes 5 and 7 A in single environments. Although not declared significant due to marginal LOD values, the QTL for FDK on the 5 and 7 A were showing in other years suggesting their effects were real. DT696 contributed the favourable alleles for low DON and FDK on all the chromosomes. Although no resistance loci contributed by DT707, transgressive segregant lines were identified resulting in greater resistance than DT696. Breeder-friendly KASP markers were developed for two of the DON and FDK QTL detected on chromosomes 5 and 7 A. Markers flanking each QTL were physically mapped against the durum wheat reference sequence and candidate genes which might be involved in FDK and DON resistance were identified within the QTL intervals. CONCLUSIONS: The DH lines harboring the desired resistance QTL will serve as useful resources in breeding for FDK and DON resistance in durum wheat. Furthermore, breeder-friendly KASP markers developed during this study will be useful for the selection of durum wheat varieties with low FDK and DON levels in durum wheat breeding programs.

A comparison between the role of enniatins and deoxynivalenol in Fusarium virulence on different tissues of common wheat.

BACKGROUND: Fusarium graminearum and Fusarium avenaceum are two of the most important causal agents of Fusarium head blight (FHB) of wheat. They can produce mycotoxins that accumulate in infected wheat heads, including deoxynivalenol (DON) and enniatins (ENNs), produced by F. graminearum and F. avenaceum, respectively. While the role of DON as a virulence factor in F. graminearum toward wheat is well known, ENNs in F. avenaceum has been poorly explored. Results obtained to-date indicate that ENNs may confer an advantage to F. avenaceum only on particular hosts. RESULTS: In this study, with the use of ENN-producing and ENN non-producing F. avenaceum strains, the role of ENNs on F. avenaceum virulence was investigated on the root, stem base and head of common wheat, and compared with the role of DON, using DON-producing and DON non-producing F. graminearum strains. The DON-producing F. graminearum strain showed a significantly higher ability to cause symptoms and colonise each of the tested tissues than the non-producing strain. On the other hand, the ability to produce ENNs increased initial symptoms of the disease and fungal biomass accumulation, measured by qPCR, only in wheat heads, and not in roots or stem bases. LC-MS/MS analysis was used to confirm the presence of ENNs and DON in the different strains, and results, both in vitro and in wheat heads, were consistent with the genetics of each strain. CONCLUSION: While the key role of DON on F. graminearum virulence towards three different wheat tissues was noticeable, ENNs seemed to have a role only in influencing F. avenaceum virulence on common wheat heads probably due to an initial delay in the appearance of symptoms.

QTL analysis of native Fusarium head blight and deoxynivalenol resistance in 'D8006W'/'Superior', soft white winter wheat population.

BACKGROUND: Fusarium head blight (FHB), caused by Fusarium graminearum, is a major disease of wheat in North America. FHB infection causes fusarium damaged kernels (FDKs), accumulation of deoxynivalenol (DON) in the grain, and a reduction in quality and grain yield. Inheritance of FHB resistance is complex and involves multiple genes. The objective of this research was to identify QTL associated with native FHB and DON resistance in a 'D8006W'/'Superior', soft white winter wheat population. RESULTS: Phenotyping was conducted in replicated FHB field disease nurseries across multiple environments and included assessments of morphological and FHB related traits. Parental lines had moderate FHB resistance, however, the population showed transgressive segregation. A 1913.2 cM linkage map for the population was developed with SNP markers from the wheat 90 K Infinium iSelect SNP array. QTL analysis detected major FHB resistance QTL on chromosomes 2D, 4B, 5A, and 7A across multiple environments, with resistance from both parents. Trait specific unique QTL were detected on chromosomes 1A (visual traits), 5D (FDK), 6B (FDK and DON), and 7D (DON). The plant height and days to anthesis QTL on chromosome 2D coincided with Ppd-D1 and were linked with FHB traits. The plant height QTL on chromosome 4B was also linked with FHB traits; however, the Rht-B1 locus did not segregate in the population. CONCLUSIONS: This study identified several QTL, including on chromosome 2D linked with Ppd-D1, for FHB resistance in a native winter wheat germplasm.

Expression of endogenous UDP-glucosyltransferase in endophyte Phomopsis liquidambaris reduces deoxynivalenol contamination in wheat.

Fusarium head blight is a devastating disease that causes severe yield loses and mycotoxin contamination in wheat grain. Additionally, balancing the trade-off between wheat production and disease resistance has proved challenging. This study aimed to expand the genetic tools of the endophyte Phomopsis liquidambaris against Fusarium graminearum. Specifically, we engineered a UDP-glucosyltransferase-expressing P. liquidambaris strain (PL-UGT) using ADE1 as a selection marker and obtained a deletion mutant using an inducible promoter that drives Cas9 expression. Our PL-UGT strain converted deoxynivalenol (DON) into DON-3-G in vitro at a rate of 71.4 % after 36 h. DON inactivation can be used to confer tolerance in planta. Wheat seedlings inoculated with endophytic strain PL-UGT showed improved growth compared with those inoculated with wildtype P. liquidambaris. Strain PL-UGT inhibited the growth of Fusarium graminearum and reduced infection rate to 15.7 %. Consistent with this finding, DON levels in wheat grains decreased from 14.25 to 0.56 µg/g when the flowers were pre-inoculated with PL-UGT and then infected with F. graminearum. The expression of UGT in P. liquidambaris was nontoxic and did not inhibit plant growth. Endophytes do not enter the seeds nor induce plant disease, thereby representing a novel approach to fungal disease control.

Detoxifying bacterial genes for deoxynivalenol epimerization confer durable resistance to Fusarium head blight in wheat.

Fusarium head blight (FHB) and the presence of mycotoxin deoxynivalenol (DON) pose serious threats to wheat production and food safety worldwide. DON, as a virulence factor, is crucial for the spread of FHB pathogens on plants. However, germplasm resources that are naturally resistant to DON and DON-producing FHB pathogens are inadequate in plants. Here, detoxifying bacteria genes responsible for DON epimerization were used to enhance the resistance of wheat to mycotoxin DON and FHB pathogens. We characterized the complete pathway and molecular basis leading to the thorough detoxification of DON via epimerization through two sequential reactions in the detoxifying bacterium Devosia sp. D6-9. Epimerization efficiently eliminates the phytotoxicity of DON and neutralizes the effects of DON as a virulence factor. Notably, co-expressing of the genes encoding quinoprotein dehydrogenase (QDDH) for DON oxidation in the first reaction step, and aldo-keto reductase AKR13B2 for 3-keto-DON reduction in the second reaction step significantly reduced the accumulation of DON as virulence factor in wheat after the infection of pathogenic Fusarium, and accordingly conferred increased disease resistance to FHB by restricting the spread of pathogenic Fusarium in the transgenic plants. Stable and improved resistance was observed in greenhouse and field conditions over multiple generations. This successful approach presents a promising avenue for enhancing FHB resistance in crops and reducing mycotoxin contents in grains through detoxification of the virulence factor DON by exogenous resistance genes from microbes.

Diametral influence of deoxynivalenol (DON) and deepoxy-deoxynivalenol (DOM-1) on the growth of Campylobacter jejuni with consequences on the bacterial transcriptome.

BACKGROUND: Deoxynivalenol (DON) is a type B trichothecene mycotoxin that is commonly found in cereals and grains worldwide. The presence of this fungal secondary-metabolite raises public-health concerns at both the agriculture and food industry level. Recently, we have shown that DON has a negative impact on gut integrity, a feature also noticed for Campylobacter (C.) jejuni. We further demonstrated that DON increased the load of C. jejuni in the gut and inner organs. In contrast, feeding the less toxic DON metabolite deepoxy-deoxynivalenol (DOM-1) to broilers reduced the Campylobacter load in vivo. Consequently, it can be hypothesized that DON and DOM-1 have a direct effect on the growth profile of C. jejuni. The aim of the present study was to further resolve the nature of this interaction in vitro by co-incubation and RNA-sequencing. RESULTS: The co-incubation of C. jejuni with DON resulted in significantly higher bacterial growth rates from 30 h of incubation onwards. On the contrary, the co-incubation of C. jejuni with DOM-1 reduced the CFU counts, indicating that this DON metabolite might contribute to reduce the burden of C. jejuni in birds, altogether confirming in vivo data. Furthermore, the transcriptomic profile of C. jejuni following incubation with either DON or DOM-1 differed. Co-incubation of C. jejuni with DON significantly increased the expression of multiple genes which are critical for Campylobacter growth, particularly members of the Flagella gene family, frr (ribosome-recycling factor), PBP2 futA-like (Fe3+ periplasmic binding family) and PotA (ATP-binding subunit). Flagella are responsible for motility, biofilm formation and host colonization, which may explain the high Campylobacter load in the gut of DON-fed broiler chickens. On the contrary, DOM-1 downregulated the Flagella gene family and upregulated ribosomal proteins. CONCLUSION: The results highlight the adaptive mechanisms involved in the transcriptional response of C. jejuni to DON and its metabolite DOM-1, based on the following effects: (a) ribosomal proteins; (b) flagellar proteins; (c) engagement of different metabolic pathways. The results provide insight into the response of an important intestinal microbial pathogen against DON and lead to a better understanding of the luminal or environmental acclimation mechanisms in chickens.

Impacts of intake of trichothecenes (Fusarium sporotrichioides) for dairy calves: Effects on animal growth, oxidative and inflammatory response.

The objective of the present study was to evaluate the impacts of trichothecenes (Fusarium sporotrichioides) for dairy calves on animal growth, oxidative and inflammatory responses in the presence or absence of essential oils. Twelve calves weaned at 70 days of age were divided into 2 groups: T-C (control) and T-EO (essential oils - oregano, thyme, basil and rosemary) in the period of 40 days consuming ration contaminated by trichothecenes (500 ppb). The animals in the T-EO group received a mixture of EOs via feed at a dosage of 0.75 mL per/kg of feed. Blood collections were performed on days 1, 20 and 40 for hematological and biochemical analyses; the fecal score was performed every 2 days on a scale of 1-5 and clinical examinations were performed 3 times during the experiment period. The animals were weighed at the beginning and at the end of the experiment; euthanasia of two calves per group for macroscopic and microscopic evaluation of several tissues (spleen, liver, duodenum, jejunum, ilium, cecum and colon) was performed at the end of the experiment. The calves in the T-EO group had a tendency (P = 0.07) of higher body weight when compared to the T-C. Treatment effect and treatment vs day interaction was detected for leukocytes and granulocytes variables, demonstrating a higher count of these cells in the T-EO group on both days (20 and 40), and the same behavior occurred for the distribution amplitude of erythrocytes (RDW). The enzymes alanine transferase (ALT), aspartate transferase (AST) and gamma glutamyl-transferase (GGT) showed higher serum activity in the T-C group (days 20 and 40). The levels of thiobarbituric acid reactive substances (TBARS) were lower in the serum of animals in the T-EO group. For calves in the T-EO group, glutathione S-transferase activity was higher in serum. Haptoglobulin and C-reactive protein levels were lower on days 20 and 40 in T-EO animals when compared to the T-C group. In the macroscopic and microscopic evaluations, which were collected at the end of the experiment after slaughtering the animals, liver and intestine did not show changes for the animals in the T-EO group, unlike the animals in the T-C group, which had moderately firm diffuse consistency of the liver and edema in the mesentery, as well as oxidative stress in tissues (liver, duodenum, jejunum, ileum, cecum and colon). The results concluded that the consumption of a mixture of EOs (essential oils - oregano, thyme, basil and rosemary) minimized the negative effects caused by trichothecenes in dairy calves, thus being an alternative to improving the immunological and antioxidant condition, as well as a possible adsorbent alternative.

Smartphone-enabled colorimetric immunoassay for deoxynivalenol based on Mn2+-mediated aggregation of AuNPs.

Deoxynivalenol (DON) is a common mycotoxin in food that mainly pollutes grain crops and feeds, such as barley, wheat and corn. DON has caused widespread concern in the field of food and feed safety. In this study, a colorimetric immunoassay was proposed based on the aggregation of gold nanoparticles (AuNPs) due to the decomposition of Mn2+ from gold-coated manganese dioxide (AuNP@MnO2) nanosheets. In this study, 2-(dihydrogen phosphate)-l-ascorbic acid (AAP) was hydrolyzed by alkaline phosphatase (ALP) and converted to ascorbic acid (AA). Then, AuNP@MnO2 was reduced to Mn2+ and AuNPs aggregation occurred. Using the unique optical characteristics of AuNPs and AuNP@MnO2, visible color changes realized simple detection of DON with high sensitivity and portability. With increasing DON content, the color changed more obviously. To quantitatively detect DON, pictures can be taken and the blue value can be read by a smartphone. The detection limit (Ic10) of this method was 0.098 ng mL-1, which was 326 times higher than that of traditional competitive ELISA, and the detection range was 0.177-6.073 ng mL-1. This method exhibited high specificity with no cross-reaction in other structural analogs. The average recovery rate of DON in corn flour samples was 89.1 %-110.2 %, demonstrating the high accuracy and stability of this assay in actual sample detection. Therefore, the colorimetric immunoassay can be used for DON-related food safety monitoring.

Deoxynivalenol Induces Drp-1-Mediated Mitochondrial Dysfunction via Elevating Oxidative Stress.

Mitochondrial dysfunction is often linked to neurotoxicity and neurological diseases and stems from oxidative stress, yet effective therapies are lacking. Deoxynivalenol (DON or vomitoxin) is one of the most common and hazardous type-B trichothecene mycotoxins, which contaminates crops used for food and animal feed. Despite the abundance of preliminary reports, comprehensive investigations are scarce to explore the relationship between these fungal metabolites and neurodegenerative disorders. The present study aimed to elucidate the precise role of DON in mitochondrial dynamics and cell death in neuronal cells. Excessive mitochondrial fission is associated with the pathology of several neurodegenerative diseases. Human SH-SY5Y cells were treated with different concentrations of DON (250-1000 ng/mL). Post 24 and 48 h DON treatment, the indexes were measured as follows: generation of reactive oxygen species (ROS), ATP levels, mitochondrial membrane potential, calcium levels, and cytotoxicity in SH-SY5Y cells. The results showed that cytotoxicity, intracellular calcium levels, and ROS in the DON-treated group increased, while the ATP levels and mitochondrial membrane potential decreased in a dose-dependent manner. With increasing DON concentrations, the expression levels of P-Drp-1, mitochondrial fission proteins Mff, and Fis-1 were elevated with reduced activities of MFN1, MFN2, and OPA1, further resulting in an increased expression of autophagic marker LC3 and beclin-1. The reciprocal relationship between mitochondrial damage and ROS generation is evident as ROS can instigate structural and functional deficiencies within the mitochondria. Consequently, the impaired mitochondria facilitate the release of ROS, thereby intensifying the cycle of damage and exacerbating the overall process. Using specific hydroxyl, superoxide inhibitors, and calcium chelators, our study confirmed that ROS and Ca2+-mediated signaling pathways played essential roles in DON-induced Drp1 phosphorylation. Therefore, ROS and mitochondrial fission inhibitors could provide critical research tools for drug development in mycotoxin-induced neurodegenerative diseases.

Identification and characterization of Achromobacter spanius P-9 and elucidation of its deoxynivalenol-degrading potential.

Deoxynivalenol (DON) poses significant challenges due to its frequent contamination of grains and associated products. Microbial strategies for mitigating DON toxicity showed application potential. Eight bacterial isolates with DON degradation activity over 5% were obtained from various samples of organic fertilizer in this study. One of the isolates emerged as a standout, demonstrating a substantial degradation capability, achieving a 99.21% reduction in DON levels. This isolate, underwent thorough morphological, biochemical, and molecular characterization to confirm its identity, and was identified as a new strain of Achromobacter spanius P-9. Subsequent evaluations revealed that the strain P-9 retains its degradation activity after a 24-h incubation, reaching optimal performance at 35 °C with a pH of 8.0. Further studies indicated that Ca2+ ions enhance the degradation process, whereas Zn2+ ions exert an inhibitory effect. This is the pioneering report of DON degradation by Achromobacter spanius, illuminating its prospective utility in addressing DON contamination challenges.

A Novel SERS Label-Free Sensing Strategy for DON and NIV: A DFT Study on the Interaction between DON/NIV and Ag/Au.

In this study, we propose a novel label-free detection strategy based on surface-enhanced Raman spectroscopy (SERS) for detecting deoxynivalenol (DON) and nivalenol (NIV), analyzing the interactions between these fungal toxins and Ag/Au substrates using density functional theory (DFT). The DFT calculation results indicate that the oxygen atoms in DON and NIV molecules exhibit high electronegativity, suggesting that they can serve as active sites in interaction with the metal surfaces in the SERS effect. By constructing DON/NIV-Ag6/Au6 complex models and evaluating their binding energies, we demonstrate the formation of stable complexes, showing characteristic frequency shifts, broadening, and selective enhancement in theoretical Raman spectra. The results indicate that the structural differences between DON and NIV lead to significant variations in the characteristic frequencies of their theoretical SERS spectra, providing a reproducible and interference-resistant method for the label-free detection of these fungal toxins. This work enhances the understanding of molecular-substrate interactions in SERS effects and provides a theoretical basis for the application of label-free SERS sensing strategies based on characteristic frequency shifts.

Methyl jasmonate mitigates Fusarium graminearum infection in wheat by inhibiting deoxynivalenol synthesis.

Methyl jasmonate (MeJA), a plant growth regulator, coordinates a diverse array of physiological responses, including the inhibition of seed germination, modulation of secondary metabolite biosynthesis, and activation of defence responses. The external application of MeJA has been demonstrated to effectively diminish the severity of fungal diseases. Here, we unveil a novel mechanism through which exogenous MeJA alleviates Fusarium head blight (FHB) by inhibiting the synthesis of deoxynivalenol (DON) in Fusarium graminearum, rather than by enhancing the wheat resistance response. MeJA treatment reduced the infection by wild-type F. graminearum in wheat coleoptiles, but exhibited no significant influence on that of the DON-deficient mutant strain (∆Tri5). The production of DON in F. graminearum was significantly inhibited both in vitro and in planta. MeJA affected the expression of genes related to DON biosynthesis, without influencing the formation of toxisomes as observed under microscopic analysis. Exogenous MeJA demonstrated a limited impact on the early genes of plant jasmonic acid signalling pathway, in contrast to the wild-type pathogen strain, which induced the upregulation of these genes. The expression levels of defence marker genes induced by MeJA were notably lower compared to those induced by the pathogen. This study elucidates the molecular mechanisms of MeJA in modulating the wheat-F. graminearum interaction, providing new insights into the development of environmentally friendly strategies against fungi.

Deoxynivalenol induces m6A-mediated upregulation of p21 and growth arrest of mouse hippocampal neuron cells in vitro.

Hippocampal neurons maintain the ability of proliferation throughout life to support neurogenesis. Deoxynivalenol (DON) is a mycotoxin that exhibits brain toxicity, yet whether and how DON affects hippocampal neurogenesis remains unknown. Here, we use mouse hippocampal neuron cells (HT-22) as a model to illustrate the effects of DON on neuron proliferation and to explore underlying mechanisms. DON exposure significantly inhibits the proliferation of HT-22 cells, which is associated with an up-regulation of cell cycle inhibitor p21 at both mRNA and protein levels. Global and site-specific m6A methylation levels on the 3'UTR of p21 mRNA are significantly increased in response to DON treatment, whereas inhibition of m6A hypermethylation significantly alleviates DON-induced cell cycle arrest. Further mechanistic studies indicate that the m6A readers YTHDF1 and IGF2BP1 are responsible for m6A-mediated increase in p21 mRNA stability. Meanwhile, 3'UTR of E3 ubiquitin ligase TRIM21 mRNA is also m6A hypermethylated, and another m6A reader YTHDF2 binds to the m6A sites, leading to decreased TRIM21 mRNA stability. Consequently, TRIM21 suppression impairs ubiquitin-mediated p21 protein degradation. Taken together, m6A-mediated upregulation of p21, at both post-transcriptional and post-translational levels, contributes to DON-induced inhibition of hippocampal neuron proliferation. These results may provide new insights for epigenetic therapy of neurodegenerative diseases.

Macrophage MAPK7/AhR/STAT3 Signaling Mediates Mitochondrial ROS Burst and Enterohepatic Inflammatory Responses Induced by Deoxynivalenol Relevant to Low-Dose Exposure in Children.

Deoxynivalenol (DON) can induce endoplasmic reticulum (ER) stress, mitochondrial ROS burst, and macrophage polarization. Here, we investigated the mechanism linking the above three aspects with the dose range relevant to low-level exposure in children. At 0.5 µg/kg bw/day, we found remarkable liver and gut inflammatory responses after 6-week exposure in mice age comparable to humans 7-12 years old. Through antioxidant intervention, we found that ROS played a driver role in macrophage polarization and inflammatory responses induced by DON in the liver and gut. Further bioinformatics analysis uncovered that ER stress-associated protein MAPK7 (ERK5) may bind with AhR to initiate a mitochondrial ROS burst and macrophage M1 polarization. The downstream cellular events of MAPK7-AhR interaction may be mediated by the AhR/STAT3/p-STAT(Ser727) pathway. This mechanism was further supported by DON toxicity mitigation using cyanidin-3-glucoside (C-3-G), which docks to MAPK7 oligomerization region 200-400 aa and disrupts MAPK7-AhR interaction. Overall, our study provides novel evidence and mechanism for DON-induced inflammatory responses in the liver and gut system. Our findings call attention to the health risks associated with low-level DON exposure in the prepuberty children population.

Eco-friendly one-step egg white gel preparation for sensitive detection of 13 trichothecenes in oats using UHPLC-MS/MS.

Oat products have gained widespread recognition as a health food due to their rich and balanced nutritional profile and convenience. However, the unique matrix composition of oats, which differs significantly from other cereals, presents specific challenges for mycotoxin analysis. This study presents an ultrahigh-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method enhanced with an innovative egg white gel pretreatment for the simultaneous analysis of 13 regulated and unregulated trichothecenes in oats. The method demonstrated excellent performance with high accuracy (> 87.5%), repeatability (< 5.7%), and reproducibility (< 8.1%). Analysis of 100 commercial oat products revealed a concerning detection rate (78%) for at least one of the 11 trichothecenes investigated. Notably, deoxynivalenol, exceeding the standard limit in 2% of samples, exhibited the highest detection rate (62%). Additionally, concerning co-occurrence patterns and positive correlations were observed, highlighting potential synergistic effects. The first-time detection of unregulated mycotoxins (T-2 triol, 4,15-diacetoxyscirpenol, 15-acetoxyscirpenol, and neosolaniol) underscores the need for comprehensive monitoring. This method, while developed for oats, shows potential for broader application to other cereals, though further investigation and confirmation are necessary. These findings suggest a potentially underestimated risk of trichothecenes in oats, necessitating continuous monitoring to ensure consumer safety.

Development of alkaline phosphatase-linked single-chain variable fragment fusion proteins for one-step immunodetection of deoxynivalenol in cereals.

Deoxynivalenol (DON) is a mycotoxin that widely distributes in various foods and seriously threatens food safety. To minimize the consumers' dietary exposure to DON, there is an urgent demand for developing rapid and sensitive detection methods for DON in food. In this study, a bifunctional single-chain variable fragment (scFv) linked alkaline phosphatase (ALP) fusion protein was developed for rapid and sensitive detection of deoxynivalenol (DON). The scFv gene was chemically synthesized and cloned into the expression vector pET25b containing the ALP gene by homologous recombination. The prokaryotic expression, purification, and activity analysis of fusion proteins (scFv-ALP and ALP-scFv) were well characterized and performed. The interactions between scFv and DON were investigated by computer-assisted simulation, which included hydrogen bonds, hydrophobic interactions, and van der Waals forces. The scFv-ALP which showed better bifunctional activity was selected for developing a direct competitive enzyme-linked immunosorbent assay (dc-ELISA) for DON in cereals. The dc-ELISA takes 90 min for one test and exhibits a half inhibitory concentration (IC50) of 11.72 ng/mL, of which the IC50 was 3.08-fold lower than that of the scFv-based dc-ELISA. The developed method showed high selectivity for DON, and good accuracy was obtained from the spike experiments. Furthermore, the detection results of actual cereal samples analyzed by the method correlated well with that determined by high-performance liquid chromatography (R2=0.97165). These results indicated that the scFv-ALP is a promising bifunctional probe for developing the one-step colorimetric immunoassay, providing a new strategy for rapid and sensitive detection of DON in cereals.

In Vitro Antimalarial Activity of Trichothecenes against Liver and Blood Stages of Plasmodium Species.

Trichothecenes (TCNs) are a large group of tricyclic sesquiterpenoid mycotoxins that have intriguing structural features and remarkable biological activities. Herein, we focused on three TCNs (anguidine, verrucarin A, and verrucarol) and their ability to target both the blood and liver stages of Plasmodium species, the parasite responsible for malaria. Anguidine and verrucarin A were found to be highly effective against the blood and liver stages of malaria, while verrucarol had no effect at the highest concentration tested. However, these compounds were also found to be cytotoxic and, thus, not selective, making them unsuitable for drug development. Nonetheless, they could be useful as chemical probes for protein synthesis inhibitors due to their direct impact on parasite synthesis processes.