Fumonisin B1 Research Articles

Mycotoxin Challenge in Dairy Cows: Assessment of the Efficacy of an Anti-Mycotoxin Agent by Adopting an In Vitro Rumen Simulation Method

To protect ruminants from the harmful effects of mycotoxins, anti-mycotoxin agents can be added to the dietary ration, thus guaranteeing animal health and production. Therefore, the objective of this study was to evaluate the in vitro ruminal initial sequestration (weak binding) and subsequent desorption (strong binding) of an anti-mycotoxin agent based on a mixture of adsorbing material, turmeric and milk thistle extracts and yeast-based components to adsorb or bio-convert aflatoxins (AF), fumonisins B1 and B2 (FB), trichothecene deoxynivalenol (DON), T-2 and HT-2 toxins, and zearalenone (ZEN). Two doses were tested: Dose 1 simulated 30 mg/cow/d, while Dose 2 simulated 90 mg/cow/d of the anti-mycotoxin agent. Each treatment involved three analytical replicates at each of three incubation times (1, 4, and 24 h post-incubation), with two independent experimental runs providing experimental replicates. Analytical methods, including UHPLC-HRMS and multivariate analyses, were used to both quantify mycotoxin concentrations and reveal dose-dependent reductions, with statistical validations indicating significant changes in mycotoxin levels across both dose and time. The results indicated that the anti-mycotoxin agent was able to highly bind AFB1, T2, and HT-2 toxins since its concentration was always under the limit of detection (<1 ppb). Regarding ZEN (weak binding mean: 94.6%; strong binding mean: 62.4%) and FBs (weak binding mean: 58.7%; strong binding mean: 32.3%), orthogonal contrasts indicated that the anti-mycotoxin agent was able to effectively bind these toxins using Dose 1 (p < 0.05). This finding suggests that Dose 1 may be sufficient to achieve the targeted effect and that a further increase does not significantly improve the outcome. Regarding DON, a strong linear relationship was observed between dose and adsorption. However, the complex interactions between the mycotoxin, the ruminal environment, and the anti-mycotoxin agent made it difficult to establish a clear dose–effect relationship (p > 0.10). UHPLC-HRMS analysis identified over 1500 mass features in rumen samples, which were further analyzed to assess the effects of the anti-mycotoxin agent. Hierarchical clustering analysis (HCA) revealed significant changes in the untargeted metabolomic profiles of samples treated with mycotoxins compared to control samples, particularly after 24 h with the anti-mycotoxin treatments. Clear differences were noted between strong binding and weak binding samples. Further analysis using orthogonal partial least squares discriminant analysis (OPLS-DA) highlighted distinct metabolomic profiles, with stronger predictive ability in the strong binding group (Q2 cumulative value of 0.57) compared to the weak binding group (0.30). The analysis identified 44 discriminant compounds in the strong binding model and 16 in the weak binding model. Seven compounds were common to both groups, while silibinin, known for its antioxidant and anti-inflammatory properties, was found among the unique compounds in the weak binding group. Overall, the findings suggest that both doses of the anti-mycotoxin agent significantly influenced the chemical profiles in the rumen, particularly enhancing the binding of mycotoxins, thereby supporting the role of phytogenic extracts in mitigating mycotoxin effects.

Structural basis of the mechanism and inhibition of a human ceramide synthase

Abstract Ceramides are bioactive sphingolipids crucial for regulating cellular metabolism. Ceramides and dihydroceramides are synthesized by six ceramide synthase (CerS) enzymes, each with specificity for different acyl-CoA substrates. Ceramide with a 16-carbon acyl chain (C16 ceramide) has been implicated in obesity, insulin resistance and liver disease and the C16 ceramide-synthesizing CerS6 is regarded as an attractive drug target for obesity-associated disease. Despite their importance, the molecular mechanism underlying ceramide synthesis by CerS enzymes remains poorly understood. Here we report cryo-electron microscopy structures of human CerS6, capturing covalent intermediate and product-bound states. These structures, along with biochemical characterization, reveal that CerS catalysis proceeds through a ping-pong reaction mechanism involving a covalent acyl–enzyme intermediate. Notably, the product-bound structure was obtained upon reaction with the mycotoxin fumonisin B1, yielding insights into its inhibition of CerS. These results provide a framework for understanding CerS function, selectivity and inhibition and open routes for future drug discovery.

The High-Efficiency Degradation of Multiple Mycotoxins by Lac-W Laccase in the Presence of Mediators

Mycotoxin cocontamination is a severe threat to health and economic security worldwide. The mycotoxins aflatoxin B1 (AFB1), zearalenone (ZEN), deoxynivalenol, T-2 toxin, fumonisin B1, and ochratoxin A are of particular concern due to their substantial toxicity. Lac-W is a laccase with the unique property of degrading these six mycotoxins in the absence of redox mediators. Nevertheless, their degradation rates are low. This work aims to improve the ability of Lac-W to degrade these six mycotoxins and to elucidate its detoxification mechanism. Including redox mediators increased the Lac-W degradation efficiency drastically, and completely degraded AFB1 and ZEN within one hour. Additionally, Lac-W-AS has good temperature, pH, and ions adaptability in ZEN degradation. Lac-W-AS reduced the ZEN toxicity because ZEN degradation products significantly restored the bioluminescence intensity of Vibrio fischeri. A Lac-W-AS-mediated oxidation product of ZEN was structurally characterized as 15-OH-ZEN by UHPLC-MS/MS. Linear sweep voltammetry showed that AS affected the potential of Lac-W and accelerated the oxidation of ZEN. Finally, the combination of mediators (acetosyringone and 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonate)) improved the degradation rate of mycotoxins. This work highlights that the combination of Lac-W with mediators serves as a good candidate for degrading multi-mycotoxin contaminants in food and feedstuff.

Toxicity of Low-Level Multiple-Mycotoxin Mixture in Nile Tilapia (Oreochromis niloticus) Is Prevented with Organically Modified Clinoptilolite Feed Additive

Organically modified clinoptilolite (member of the zeolite family of minerals; MinazelPlus®) feed additive, with an average weight of 30 ± 2 g, was used to prevent mycotoxicosis in Nile tilapia (Oreochromis niloticus) through its supplementation for 42 days to the diet contaminated with multiple mycotoxins: aflatoxin B1 (40 µg/kg), fumonisin B1 and B2 (600 µg/kg), zearalenone (50 µg/kg), and deoxynivalenol (150 µg/kg). The fish were divided randomly into four experimental groups (basal diet control—C; fed 2 g/kg MinazelPlus®—MZ; fed multiple mycotoxins—MT; and fed a combination of MinazelPlus® and multiple mycotoxins—MZ + MT). Each group consisted of triplicate aquarium setups, with six fish in each replicate. Sampling was performed in weeks 2, 4, and 6. The lymphocyte count was significantly higher in the MZ group compared with the MT group and the MT + MZ group in week 6. An overall decrease in the neutrophil count was observed in the experimental groups. Histopathological analysis was performed in weeks 2 and 6, revealing significant changes in the liver, intestines, kidney, and spleen of fish from the MT group, while the MT + MZ and MZ groups were similar to the control. The addition of 2 g/kg MinazelPlus® has the ability to prevent and reduce the adverse effects of chronic exposure to low concentrations of multiple mycotoxins in juvenile Nile tilapia.

Determination of some mycotoxins from poultry feed in Baghdad city

The aims of this study were to determine some mycotoxins and to isolate the fungi from chicken feed samples from broiler and layer farms in Baghdad city. The isolation rate (percentage) of fungi from poultry feed samples was Aspergillus niger 17.7%, Aspergillus flavus 14.7%, Aspergillus fumigatus 12.4%, Aspergillus ochraceus 10.7%, Penicillium spp. 9.3%, Fusarium spp. 9.3%, Rhizopus spp. 8.4%, Mucar spp. 8.4%, Absidia spp. 7.6% and Chrysonilia sitophila 1.5%, while the most common yeast genera were Candida spp. 44.8%, Candida albicans 20.7%, Cryptococcus neoformans 17.3%, Geotricum candidum 12% and Trichosorom spp. 5.2%. The mycotoxins ochratoxin A (OTA), deoxynivalenol (DON), fumonisin B1 (FB1) and zearalenone (ZON) were detected, and the percentages of these mycotoxins were 43.1%, 27.2%, 21.1% and 8.6%, respectively. This study indicates that the presence of mold that produce mycotoxins in poultry feed increases the risk of mycotoxins in feed, meat, eggs, etc. of animals and therefore humans, and we need to raise awareness of the Ministry of Agriculture and Health to prevent these measures to reduce the mycotoxin levels in poultry products.

Unveiling the Substrate-Dependent Dynamics of Mycotoxin Production in Fusarium verticillioides Using an OSMAC-Metabolomics Approach.

Fusarium verticillioides is a prevalent plant pathogenic fungus known to produce harmful mycotoxins, including fumonisins and emerging toxins. This study aimed to investigate the influence of substrate on the temporal patterns of mycotoxin biosynthesis by F. verticillioides, employing a combined OSMAC (One Strain-Many Compounds) strategy and metabolomics approach. The fungus was cultured under various media conditions, and samples were collected over time. LC-MS/MS analyses and a dereplicative workflow were used to profile the secondary metabolite production, focusing on mycotoxins. The results demonstrated that modifying the culture conditions led to significant variations in fungal growth and the nature and relative concentrations of mycotoxins produced. Corn meal agar (CMA) medium was favorable for fumonisins A1 and B1, while malt extract agar (MEA) favored fumonisins A2 and B2. The study also identified the production of other mycotoxins related compounds as fusarins, bikaverin derivatives and fumonisins analogs, under different growth conditions. This study highlights the potential of combining OSMAC and metabolomics to unravel the substrate-dependent and time-dependent variations in mycotoxin biosynthesis by F. verticillioides. The insights gained provide a better understanding of the ecophysiology of this fungus and the occurrence of its mycotoxins, which can inform targeted mitigation strategies to ensure food and feed safety.

Determination of mycotoxins in breakfast cereals by LC-MS/MS

Breakfast cereals, attributed to low microbiological risk due to their low water activity, are not very innocent due to their components such as cereal, dried fruit and cocoa. Secondary fungal metabolites called mycotoxins are one of the most important risks that these foods can carry. The multimycotoxin level of 140 breakfast cereal samples sold in local markets in Istanbul, Turkiye, was analysed.Samples were tested using the LC-MS/MS technique. Contamination levels of aflatoxin B1, B2, G1, G2, ochratoxin (OTA), zearalenone (ZEN), deoxynivalenol (DON), fumonisin B1, B2, citrinin, HT-2 and T-2 toxins were determined in breakfast cereal.As a result, 71% (n = 100) of the samples were contaminated with at least one mycotoxin. Aflatoxin B1 levels in 17% of the samples and total aflatoxin levels in 31% were determined to exceed the limits defined in the “Turkish Food Codex Contaminants Regulation”. The sample rate containing all toxins (except CIT) above the detection limit was 24%. ZEN + DON + OTA was found in 24% of the samples, total aflatoxins (TAF) + OTA + ZEN in 25%, fumonisins (FUM) + OTA + DON in 29%, and TAF + FUM + OTA + DON + ZEN + HT-2+T-2 in 24% of the samples.Our findings indicate that breakfast cereal samples are susceptible to mycotoxin contamination and necessitate consistent monitoring to ensure safety. The presence of other mycotoxins not included in the legal regulation is noteworthy. Action is needed to prepare a legal regulation on the co-occurrence of mycotoxins.

Sulcatone as a Plant-Derived Volatile Organic Compound for the Control of the Maize Weevil and Its Associated Phytopathogenic Fungi in Stored Maize.

Stored maize is frequently attacked by different pests, such as insects and microorganisms. The aim of the present study was to evaluate the bioactivities of sulcatone (6-methyl-5-hepten-2-one) against the maize weevil Sitophilus zeamais and the phytopathogenic fungi Fusarium verticillioides, Aspergillus flavus, and A. parasiticus. Sulcatone showed a strong repellent effect with a maximum value of -92.1 ± 3.2% at 40 µM in two-choice olfactometer bioassays and an LC95 value of 17.2 µL/L air (95% 16.5-18.1) in a fumigant toxicity experiment. The antifungal effect of sulcatone was evaluated through the fumigant method, reporting MIC values of 3.5, 3.8, and 3.9 mM for F. verticillioides, A. parasiticus, and A. flavus, respectively. Additionally, a silo-bag experiment containing all pests was conducted to evaluate the potential use of sulcatone in a real storage system. Sulcatone caused 71.69 ± 1.57% weevil mortality in silo-bags and proved to be effective as a fungicidal and antimycotoxigenic agent since both ergosterol and fumonisin B1 content were significantly reduced by 60% in silo-bags containing sulcatone. This study demonstrated that sulcatone has the potential to be used for the control of both insects and fungi of stored maize, without affecting the germination of grains.

The effects of fumonisin B1 on intercellular communications and miRNA modulations: Non-genotoxic carcinogenesis mechanisms in human kidney cells

Fumonisin B1 (FB1), which is produced by Fusarium species, is one of the most prevalent mycotoxins known to exert several toxic effects, particularly nephrotoxicity. While its genotoxic carcinogenic mechanisms have been extensively studied, its influence on non-genotoxic pathways including intercellular communication and microRNA (miRNA) regulation remain underexplored. The present study investigates the effects of FB1 on gap junctions, miRNA expression profiles, and their relationship in human kidney cells (HK-2 and HEK293). Both cell lines showed increased apoptosis rates at 50 and 100 µM, while FB1 exposure significantly reduced gap junctional intercellular communication (GJIC) and decreased the expression levels of related genes, including Cx43, Cx45, e-cadherin, Cadherin-2, and β-catenin. After FB1 treatments alteration on the regulation of miRNAs including let-7a-5p, miR-125a-5p, miR-222–3p, miR-92a-3p, let-7b-5p, let-7e-5p, miR-21–5p, miR-155–5p, let-7i-5p, let-7d-5p, let-7f-5p, miR-181b-5p, miR-15b-5p, miR-23b-3p, miR-20b-5p, miR-196a-5p miRNAs have been shown. Let-7a-5p was selected among the altered miRNAs to elucidate the relationship between miRNAs and GJIC after FB1 exposure as it is one of the common miRNAs that changes in both cell lines and one of its target genes is Cx45, which is an important gene for GJIC. However, transfection analysis did not show any differences, resulting in Cx45 not being a direct target of let-7a-5p in HK-2 and HEK-293 cells. Through comprehensive analysis, we elucidated that FB1's impact on intercellular signaling cascades and its regulatory role on miRNA expression profiles, offering valuable insights into carcinogenesis beyond traditional genotoxic paradigms. Understanding these mechanisms is crucial for elucidating the mechanisms of FB1-induced toxicity.

Near-infrared-driven dual-photoelectrode photoelectrochemical sensing for fumonisin B1: Integrating a photon up-conversion bio-photocathode with an enhanced light-capturing photoanode

Fumonisin B1 (FB1), the most prevalent and highly toxic mycotoxin within the fumonisins family, poses threats to humans, especially in children and infants, even at trace levels. Therefore, it is essential to design an easy and sensitive detection strategy. Herein, a brand-new dual-photoelectrode photoelectrochemical (PEC) sensing platform for FB1 detection under near-infrared irradiation was unveiled. This platform integrated a photon up-conversion bio-photocathode substrate (UCNPs/Au/CuInS2, UCNPs: NaYF4: Yb3+, Er3+, Nd3+) and used a SnO2/SnS2@Bi/Bi2S3 heterojunction photoanode to greatly enhance light capture. Additionally, ZnO coated with polydopamine (ZnO@PDA) was utilized as a signal inhibitor. The restoration of photocurrent occurred due to the strong binding affinity between FB1 and its aptamer (FB1-Apt), facilitating the dissociation of FB1-Apt/ZnO@PDA from the photoelectrode. The PEC sensing performance and the electron transfer process were thoroughly examined. The developed “signal-restoration” PEC aptasensor exhibited a wider dynamic linear range from 1.0 × 10−3 to 1.0 × 102 ng/mL, with a lower limit of detection (0.13 pg/mL). It has demonstrated excellent practical detection performance in unspiked real samples, such as corn paste, with the FB1 enzyme-linked immunosorbent assay (ELISA) Kit serving as a reference, indicating its potential for routine analysis of other mycotoxins. Thus, this research establishes a feasible dual-photoelectrode PEC framework for the effective detection of mycotoxins and other hazardous substances.

Assessment of Mycotoxins in Infants and Children Cereal-Based Foods: Dietary Exposure and Potential Health Risks

AbstractPublic health concerns are rising regarding potential mycotoxin contamination in processed cereal-based products consumed by infants and young children. Due to their developing physiological systems and greater vulnerability, children are particularly susceptible to the adverse health effects of mycotoxins. This study investigated the presence of a wide range of mycotoxins (20 in total, encompassing various chemical classes, such as aflatoxins, ochratoxins, trichothecenes, zearalenones, fumonisins, citrinin and sterigmatocystin) in readily available processed cereal-based foods targeted towards infants and children in Portugal. Among the 148 analysed samples, 43% contained at least one of the targeted mycotoxins. Aflatoxin B1 (8.1%) and fumonisin B1 (10.8%) were the most commonly detected regulated mycotoxins, while emerging mycotoxins like enniatin B (14.9%) and beauvericin (10.1%) were also prevalent. The most commonly co-occurring mycotoxins were ENNB + BEA and FB1 + FB2. Although the Estimated Daily Intake of non-carcinogenic mycotoxins in most products was below their respective Tolerable Daily Intake (DONs and FBs (1 μg/kg bw/day), HT-2 and T-2 (0.02 μg/kg bw/day), ZEN (0.25 μg/kg bw/day), and CIT (0.2 μg/kg bw/day)), Hazard Quotient and Hazard Index values exceeded safe thresholds, indicating potential health risks.

Volatilome of the maize phytopathogenic fungus Fusarium verticillioides: potential applications in diagnosis and biocontrol.

Fusarium verticillioides is a maize fungal phytopathogen and a producer of volatile organic compounds (VOCs) and fumonisin B1 (FB1). Our aim was to study the volatilome, conidial production, ergosterol and FB1 biosynthesis in maize cultures over a 30-day incubation period (5, 10, 15, 20, 25, 30 days post inoculation [DPI]). The effect of pure VOCs on the same parameters was then evaluated to study their potential role as biocontrol agents. In total, 91 VOCs were detected, with volatile profiles being more similar between 5 and 10 DPI compared with 15, 20, 25 and 30 DPI. Ergosterol content increased steadily with incubation time, and three growth stages were identified: a lag phase (0 to 15 DPI), an exponential phase (15 to 20 DPI) and a stationary phase (20 to 30 DPI). The maximum concentration of FB1 was detected at 25 (0.030 μg FB1/μg ergosterol) and 30 DPI (0.037 μg FB1/μg ergosterol), whereas conidial production showed a maximum value at 15 DPI (4.3 ± 0.2 × 105 conidia/μg ergosterol). Regarding pure VOCs, minimal inhibitory concentration values ranged from 0.3 mm for 4-hexen-3-one to 7.4 mm for 2-undecanone. Pure VOCs reduced radial growth, conidial production and ergosterol and FB1 biosynthesis. The marked resemblance between VOC profiles at 5 and 10 DPI suggests that they could act as early indicators of fungal contamination, particularly 4-ethylguaiacol, 4-ethyl-2-methoxyanisole, heptanol and heptyl acetate. On the other hand, their role as inhibitors of fungal growth and FB1 biosynthesis prove their great potential as safer alternatives to control phytopathogenic fungi. © 2024 Society of Chemical Industry.

Apocynin protects the dopaminergic cell line SH-SY5Y from fumonisin B1 induced cytotoxicity through attenuation of oxidative stress mediated apoptosis

BackgroundThe fusarium verticillioides produces the mycotoxin fumonisin b1 (Fb1), commonly infects corn and other agricultural commodities. The Fb1 showed hepatotoxicity, cytotoxicity, and carcinogenicity in animals. Aim of the studyThere are limited literature available on cytotoxicity exerted by Fb1. Hence, the present investigation aimed to evaluate the cytotoxic effects of Fb1 and its mechanism in the neuroblastoma (SH-SY5Y) cell line and its amelioration by apocynin. MethodsWe investigated the molecular mechanism of Fb1-induced cytotoxicity in SH-SY5Y cells and investigated the effect of Fb1 oxidative stress markers, and apoptosis. ResultsThe results explained that the Fb1 showed dose-dependent cytotoxicity in SH-SY5Y cells with the IC50 value 150 µM. Fb1 elevated reactive oxygen species and depolarized mitochondrial membrane potential, vacuolation and apoptotic changes were observed in SH-SY5Y cells. The protein expression of CAT and GPx were downregulated with Fb1 treatment, and apoptotic markers caspase-3 and caspase-8 were upregulated. When the apocynin was pretreated, followed by Fb1 exposure for 24 h, all the changes were normalized with the treatment. Hence, these results suggest that ROS is the primary upstream signal leading to increased Fb1-mediated cytotoxicity in SH-SY5Y cells. ConclusionApocynin reversed the Fb1-induced oxidative stress and apoptosis by its antioxidant potency in SH-SY5Y cell lines.

Establishment of appropriate conditions for the efficient determination of multiple mycotoxins in tea samples and assessment of their drinking risks

Conditions were determined for rapid, convenient, and efficient determination of 16 common mycotoxins in tea samples. Mycotoxins in tea leaves and tea infusion samples were extracted using solid-liquid extraction/liquid-liquid extraction combined with ultrasonic-assisted extraction. The extraction solvent was 2-butanone/ethyl acetate (9/1 v/v) with 0.1 % formic acid. The established conditions enabled the analysis of these mycotoxins by ultra-high-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) in 5.5 min. In addition, HPLC with a temperature-controlled fluorescence detector was able to simultaneously determine 8 mycotoxins with fluorescent properties in 10 min without derivatization. Aflatoxin M1 (2.15 and 3.01 μg/kg), fumonisin B2 (198.89 μg/kg), and zearalenone (87.54 μg/kg) could be detected in commercially available pu-erh tea, green tea, and black tea products, and their total transfer rates from the products to brewed tea infusions were 64.08–65.13 %, 90.59 %, and 25.99 %, respectively. The risks of drinking mycotoxins from these tea infusions mostly showed low levels of concern.

Effects of fumonisin B1 on gene-specific DNA methylation related to cancer pathways of p53 signalling and PI3K/Akt signalling in human kidney cells

Effects of fumonisin B1 on gene-specific DNA methylation related to cancer pathways of p53 signalling and PI3K/Akt signalling in human kidney cells

Regulation of Fumonisin B1 Production and Pathogenicity in Fusarium verticillioides by Histone Deacetylases

Transcriptional regulation mediated by the balance of histone acetylation and deacetylation is fundamental in responding to environmental cues by impacting chromatin remodeling. Histone deacetylases (HDACs) are enzymes that remove acetyl groups from histone and non-histone proteins, thus restoring a tight chromatin structure. In pathogenic fungi, HDACs have been implicated in growth, secondary metabolite biosynthesis, and virulence. However, the role of HDACs in the mycotoxin fumonisin B1 (FB1)-producing fungus Fusarium verticillioides is poorly understood. In this study, we systematically characterized six F. verticillioides HDACs. An increased level of H4K16ac was observed in the deletion mutant of FvHOS2, which was associated with vegetative growth, conidiation, and virulence when infecting sugarcane and maize. FvRpd3 appeared to be essential for vegetative growth, while FvHda1 promoted growth, and both contributed to conidiation and pathogenicity. In contrast, FvSirt4 displayed a negative correlation with these processes. Additionally, the FB1 production was positively affected by FvHos2 and FvRpd3, but negatively impacted by Fvhda1, FvSir2, FvHst2, and FvSirt4 through the regulation of different key fumonisin biosynthetic (FUM) genes. Further findings indicate an association between FvSirt4 and FvSkb1, which is a histone methylase that positively regulates FB1 and pathogenicity. Moreover, as a global transcriptional regulator, over 2365 genes (~15% of the genome) enriched in multiple metabolic pathways were significantly downregulated in the ΔFvhos2 mutants relative to the wild type. Overall, our results suggest distinct roles of HDACs in regulating the growth, virulence, mycotoxin FB1 production, and gene expression in F. verticillioides.

Unlocking the Potential of Lactococcus lactis subsp. lactis BIONCL17752 strain on Fumonisin B1 production by Fusarium verticillioides

Mycotoxins are the potent toxic chemical agent's linked to food safety, pose a serious risk to human and animal health globally. The management of toxigenic fungi is a challenging task in food and feed industries. The present investigation aimed to use probiotic potentials of lactic acid bacteria (LAB), and fermented cell-free broth as antifungal agents and neutralization of carcinogenic mycotoxin, fumonisin B1 (FB1) in food and feed considered as “turned waste into treasure”. We strived to investigate the probiotic potential of Lactococcus lactis subsp. lactis BIONCL17752 strain on prevention of growth and neutralization of FB1 produced by Fusarium verticillioides BIONCL4 strain to emulate as as a food grade bio-preservative. The BIONCL17752 strain exhibited excellent inhibitory activity against BIONCL4 strain. It was cultivated in MRS medium, and obtained cells pellet (CP) and cell-free supernatant (CFS) was lyophilized, and used for their antifungal activity against BIONCL4 strain. The minimum fungicidal concentration (MFC) determined as 12.5 μg/mL. Nevertheless, CFS assessed for neutralization of FB1 production which exhibited complete inhibition and downregulation of FB1 encoding FUM1 gene expression using quantitative real-time PCR (qPCR). Furthermore, CFS induced transcriptomic studies against BIONCL4 strain endorsed a significant downregulation of virulence, FB1, fusaric acid, fusarin, and chitin biosynthetic pathway genes. The CFS and cells reticent spore germination and FB1 production in the range of 40–61% and 74–85%, respectively under stored maize for 60 days. The current findings suggest BIONCL17752 strain apprehend the fungal growth and inhibit the FB1, and other toxigenic molecules biosynthesis, and can be employed as food grade preservative in the food industries to ensure the food safety and human health.

An fusaric acid-based CRISPR library screen identifies MDH2 as a broad-spectrum regulator of Fusarium toxin-induced cell death

Fusarium mycotoxins are of great concern because they are the most common food-borne mycotoxins and environmental contaminants worldwide. Fusaric acid (FA), Deoxynivalenol (DON), Zearalenone (ZEA), T-2 toxin (T-2), and Fumonisin B1 (FB1) are important Fusarium toxins contaminating feeds and food and can cause serious health problems. FA can synergize with some other Fusarium toxins to enhance overall toxicity. However, the underlying molecular mechanism remains poorly understood. In this study, our CRISPR screening revealed Malate dehydrogenase 2 (MDH2) and Pyruvate dehydrogenase E1 subunit beta (PDHB) are the key genes for FA-induced cell death. Pathways associated with mitochondrial function, notably the TCA cycle, play a significant role in FA cytotoxicity. We found that MDH2 and PDHB depletion reduced FA-induced cell death, ROS accumulation, and the expression of caspase-3 and HIF-1α. The cell viability assays and flow cytometry demonstrated that MDH2 knockout but not PDHB decreased DON, ZEA, T-2, and FB1-induced cytotoxicity, apoptosis, and ROS accumulation. MDH2 inhibitor LW6 also decreased DON, ZEA, T-2, and FB1-induced toxicity. This suggested that MDH2, but not PDHB, is a common regulator of broad-spectrum Fusarium toxin (FA, DON, ZEA, T-2, and FB1)-induced cell death. Our work provides new avenues for the treatment of Fusarium toxin toxicity.

First evidence on the occurrence of multi-mycotoxins and dietary risk exposure to AFB1 along the cassava value chain in Uganda

This study investigated the occurrence and distribution of multiple mycotoxins (aflatoxin B1, B2, G1, G2, fumonisins B1, B2, ochratoxin A (OTA), deoxynivalenol (DON), zearalenone (ZEN), and citrinin (CIT)) in cassava products and as assessed the potential risk of aflatoxin B1 (AFB1) exposure among cassava consumers. A total of 192 samples of cassava products (96 flour and 96 chips, each with 48 samples from farmer and 48 from wholesaler) were analysed using LC/MS–MS. All positive samples irrespective of their origin (flour or chips) exhibited AFB1 levels exceeding the EU regulatory threshold of 5 µg/kg. The sum of fumonisins (FB1 + FB2), ZEN, and DON were significantly (P < 0.05) higher in cassava flour (14.3 µg/kg; 3.71 µg/kg; 25.1 µg/kg) compared to chips (6.54 µg/kg; 1.25 µg/kg; 0.25 µg/kg), respectively. Aflatoxins G2 was not detected in any of 192 samples. Cassava flour samples from farmers exhibited significantly (P < 0.05) higher mean concentrations of AFB1 (27.1 µg/kg), total aflatoxins (78.2 µg/kg), and ochratoxin A (79.6 µg/kg) in contrast to wholesalers, whose mean levels were notably lower at 8.91, 5.79 µg/kg, and 2.44 µg/kg, respectively, pointing the likely critical source of mycotoxin contamination. Cassava consumers in Northern Uganda are at a higher risk, with an estimated 2.06 cancer cases per 100,000 individuals per year compared to those in Eastern Uganda at 0.25. This study underscores the urgent need for interventions to manage aflatoxins in cassava flour, particularly at farm level in Northern Uganda. It accentuates a shift market to household-level sampling and the need for analytical methods targeting multiple mycotoxins.

Functional analysis of cutinase transcription factors in Fusarium verticillioides

Fusarium verticillioides is an important pathogen of maize and causes serious yield losses and food safety issues worldwide. F. verticillioides produces highly toxic mycotoxin Fumonisin B1 (FB1) in infested commodities which makes these food and feeds unsafe for humans and animals. For pathogenic fungi to successfully penetrate its plant hosts, the pathogen secretes hydrolytic enzymes that can facilitate penetration into the plant cutin layer. However, there is limited information on how cutinases transcriptionally regulated to impact F. verticillioides pathogenicity. In this study, our aim is to functionally characterize cutinase transcription factors that regulate key cutinase activities that are directly associated with F. verticillioides pathogenicity and FB1 biosynthesis. Gene deletion of cutinase transcription factor FvCTF1α did not affect the growth and morphology of the fungal mycelia on CMII medium, whereas the conidiation, utilization of sodium acetate and sodium oleate, stress tolerance against cell wall interfering agent, and the cutinase and pectinase activities in the ΔFvctf1α mutant were negatively impacted. FvCtf1α regulates the expression of induced cutinase genes FvCUT1 and FvCUT4 by binding to their GC-rich promoters. In addition, FvCtf1α, containing a novel function in regulating FB1, interacts with the promoter of FvFUM1 and FvFUM6 to down-regulate the expression of FvFUM1 and FvFUM6, resulting in decreased production of FB1 in the ΔFvctf1α strain. ΔFvctf1α exhibited decreased pathogenicity in maize due to the down-regulation of pathogenicity-related genes as well as key downstream cutinase genes FvCUT3 and FvCUT4 in F. verticillioides. We also demonstrated that FvCtf1α regulated FvCUT3 and FvCUT4 differently; FvCUT4 via direct regulation while FvCUT3 via indirect regulation by interacting with FvFarB, a homologous protein of FvCtf1α. Moreover, RNA-seq analysis showed that FvCtf1α was associated with many pathways, such as fatty acid metabolism, carbon source utilization, cell wall integrity, oxidative stress, and fumonisin synthesis in F. verticillioides. Our study demonstrated that FvCtf1α was not only involved in the regulation of cutinases but also a broad spectrum of pathways that ultimately affect F. verticillioides virulence and mycotoxin biosynthesis.