Occurrence of mycotoxins in meat alternatives: Dietary exposure, potential health risks, and burden of disease.

This study aimed to present the occurrence of sixteen mycotoxins in 105 meat alternatives based on wheat, legumes, and vegetables from Italy. The targeted mycotoxins were aflatoxins (AFB1, AFB2, AFG1, AFG2), fumonisins B1 and B2 (FB1, FB2), alternariol (AOH), alternariol monomethyl ether (AME), tentoxin (TEN), ochratoxin A (OTA), zearalenone (ZEN), T-2/HT-2 toxin, deoxynivalenol (DON), enniatin B (ENNB), and beauvericin (BEA). The occurrence of mycotoxins was between 0% (AFB2) - 97.4% (ENNB). Mycotoxin co-occurrence varied from binary combinations up to mixtures of twelve. To assess the dietary exposure and potential health risks we simulated the replacement of meat consumption for Italian consumers with meat alternatives. The cumulative exposure to Alternaria mycotoxins and trichothecenes indicated a potential health risk while the exposure to aflatoxins and ochratoxin A indicated a potential health concern related to liver and renal cancer in the model scenario. Moreover, we estimated the risk of liver cancer from exposure to AFB1 and quantified the potential burden using Disability-Adjusted Life Years (DALYs). Luckily, the potential risk of liver cancer was low between 0 and 0.05/100,000 individuals with an associated burden of disease of 0.83 DALYs/100,000 individuals. Taking into consideration the presence of meat alternatives on the food market and the ongoing shift towards plant-based diets there is a need for continuous monitoring to keep the occurrence at safe levels. More attention is needed from the regulatory side for policymakers to consider the legislations of mycotoxins in meat alternatives.

Correlation between metabolites of lactic acid bacteria isolated from dairy traditional fermented Tunisian products and antifungal and antioxidant activities.

AIMS: The objective of this study is to identify and investigate the antifungal and antioxidant potential of lactic acid bacteria (LAB) isolated from traditional fermented products. METHODS AND RESULTS: In this work, a collection of LAB was isolated from traditional fermented products collected in four Tunisian regions. After first screening using the overlay method, seven bacterial strains were retained due to their high antifungal effect. Four strains of Limosilactobacillus fermentum were identified, one strain of Lacticaseibacillus paracasei, one strain of Lacticaseibacillus rhamnosus and one strain of Enterococcus faecium. The antifungal and the antioxidant potential of these bacteria were then evaluated. Bacterial strains were effective against six fungal strains with minimum inhibitory concentrations ranging from 25 to 100 mg/ml and minimum fungicidal concentrations ranging from 50 to 200 mg/ml. Cell-free supernatants of LAB were analysed by HPLC-DAD and LC-MS-qTOF-MS analysis. Results showed significant production of organic acids as well as several phenolic compounds. Correlation analysis confirmed that PLA and 1,2-dihydroxybenzene were positively correlated with antifungal potential. The results of the antioxidant activity highlighted an ABTS radical cation scavenging activity ranging from 49% to 57% and a DPPH trapping percentage ranging from 80% to 97%. CONCLUSIONS: Therefore, due to these characteristics, identified lactic acid bacteria strains have shown their effectiveness to perform as antifungal and antioxidant agents. SIGNIFICANCE AND IMPACT OF THE STUDY: Since microbial contamination is at the root of extensive losses in the food sector, the identified strains or their metabolites can potentially be used as additives to limit micro-organism spoilage in food products and increase their shelf life.

Use of Mustard Extracts Fermented by Lactic Acid Bacteria to Mitigate the Production of Fumonisin B1 and B2 by Fusarium verticillioides in Corn Ears.

Corn (Zea mays) is a worldwide crop subjected to infection by toxigenic fungi such as Fusarium verticillioides during the pre-harvest stage. Fusarium contamination can lead to the synthesis of highly toxic mycotoxins, such as Fumonisin B1 (FB1) and Fumonisin B2 (FB2), which compromises human and animal health. The work aimed to study the antifungal properties of fermented yellow and oriental mustard extracts using nine lactic acid bacteria (LAB) in vitro. Moreover, a chemical characterization of the main phenolic compounds and organic acids were carried out in the extracts. The results highlighted that the yellow mustard, fermented by Lactiplantibacillus plantarum strains, avoided the growth of Fusarium spp. in vitro, showing Minimum Inhibitory Concentration (MIC) and Minimum Fungicidal Concentration (MFC) values, ranging from 7.8 to 15.6 g/L and 15.6 to 31.3 g/L, respectively. Then, the lyophilized yellow mustard fermented extract by L. plantarum TR71 was applied through spray-on corn ears contaminated with F. verticillioides to study the antimycotoxigenic activity. After 14 days of incubation, the control contained 14.71 mg/kg of FB1, while the treatment reduced the content to 1.09 mg/kg (92.6% reduction). Moreover, no FB2 was observed in the treated samples. The chemical characterization showed that lactic acid, 3-phenyllactic acid, and benzoic acid were the antifungal metabolites quantified in higher concentrations in the yellow mustard fermented extract with L. plantarum TR71. The results obtained confirmed the potential application of fermented mustard extracts as a solution to reduce the incidence of mycotoxins in corn ears.

Application of White Mustard Bran and Flour on Bread as Natural Preservative Agents.

In this study, the antifungal activity of white mustard bran (MB), a by-product of mustard (Sinapis alba) milling, and white mustard seed flour (MF) was tested against mycotoxigenic fungi in the agar diffusion method. The results obtained were posteriorly confirmed in a quantitative test, determining the minimum concentration of extract that inhibits the fungal growth (MIC) and the minimum concentration with fungicidal activity (MFC). Since MF demonstrated no antifungal activity, the MB was stored under different temperature conditions and storage time to determine its antifungal stability. Finally, an in situ assay was carried out, applying the MB as a natural ingredient into the dough to avoid P. commune CECT 20767 growth and increase the bread shelf life. The results demonstrated that the antifungal activity of MB was dose-dependent. The higher assayed dose of MB (10 g/kg) reduced the fungal population in 4.20 Log CFU/g regarding the control group. Moreover, the shelf life was extended four days compared to the control, equaling its effectiveness with the synthetic preservative sodium propionate (E-281). Therefore, MB could be an alternative to chemical additives in bread formulations since it satisfies consumer requirements. Also, the formulation of bread with MB valorizes this by-product generated during mustard seed milling, thereby helping the industry move forward sustainably by reducing environmental impact.

Development of an Antifungal Device Based on Oriental Mustard Flour to Prevent Fungal Growth and Aflatoxin B1 Production in Almonds.

The present study describes the manufacture of an antifungal device composed of oriental mustard flour and hydroxyethyl-cellulose (H-OMF) and evaluates its efficacity in inhibiting Aspergillus flavus growth and aflatoxin B1 (AFB1) production in almonds. Additionally, it compares the H-OMF with allyl isothiocyanate (AITC) and a freeze-dried extract of yellow mustard flour (YMF-E); such substances were previously described as antifungal. Minimum inhibitory concentration (MIC), Minimum fungicidal concentration (MFC), the H-OMF in vitro antifungal activity, and the residual fungal population, as well as the production of AFB1 in almonds were determined. AITC and YMF-E showed significant antifungal activity in vitro. Additionally, the in vitro activity of H-OMF avoided mycelial growth by applying 30 mg/L. Almonds treated with AITC (5.07, 10.13, and 20.26 mg/L) and H-OMF (2000 and 4000 mg/L) showed a reduction in the population of A. flavus and the production of AFB1 to values below the limit of detection. YMF-E showed effectiveness by in vitro methodologies (MIC and MFC) but did not show efficacy when applied in almonds. Our findings indicated that the hydroxyethyl-cellulose-based device containing oriental mustard flour might be utilised as a fumigant to increase the safety of almonds and could be extended to other cereals or dry fruits.

Control of Fusarium graminearum in Wheat With Mustard-Based Botanicals: From in vitro to in planta.

Fusarium graminearum is a phytopathogenic fungus that causes Fusarium head blight in small-grain cereals, such as wheat, with significant yield reductions. Moreover, it contaminates the cereal grains with health-threatening mycotoxins, such as deoxynivalenol (DON), jeopardizing food and feed safety. Plant-based biopesticides, i.e. botanicals, have recently gained increased interest in crop protection as alternatives to synthetic chemical products. The main objective of this study was to test the control efficacy of botanicals based on white or Indian/Oriental mustard seed flours (Tillecur - Ti, Pure Yellow Mustard - PYM, Pure Oriental Mustard - POM, Oriental Mustard Bran - OMB) on F. graminearum infection and mycotoxin accumulation in wheat grain. Botanicals at 2% concentration showed a higher efficacy in inhibiting mycelium growth in vitro compared with a prothioconazole fungicide (F). In the growth chamber experiment under controlled conditions, the spraying agents reduced DON content in grain in the following order: F = Ti = PYM > POM > OMB. The antifungal activity of the botanicals may be attributed to their bioactive matrices containing isothiocyanates (ITCs) and phenolic acids. Allyl ITC was detected in POM and OMB at 8.38 and 4.48 mg g-1, while p-hydroxybenzyl ITC was found in Ti and PYM at 2.56 and 2.44 mg g-1, respectively. Considerable amounts of various phenolic acids were detected in all botanicals. Under field conditions, only the use of F significantly decreased F. graminearum infection and DON content in grain. An additional important finding of this study is that disease control was more difficult when infection was done with ascospores than conidia, which might have several potential implications considering that ascospores are more important in Fusarium head blight epidemics. Our results suggest that mustard-based botanicals are promising biopesticides for the control of Fusarium head blight in small-grain cereals, but for field applications, an appropriate formulation is necessary to stabilize and prolong the antifungal activity, especially against ascospores.

Use of Botanicals to Suppress Different Stages of the Life Cycle of Fusarium graminearum.

Fusarium head blight (FHB) is one of the most important cereal diseases worldwide, causing yield losses and contamination of harvested products with mycotoxins. Fusarium graminearum is one of the most common FHB-causing species in wheat and barley cropping systems. We assessed the ability of different botanical extracts to suppress essential stages of the fungal life cycle using three strains of F. graminearum (FG0410, FG2113, and FG1145). The botanicals included aqueous extracts from white mustard (Sinapis alba) seed flour (Pure Yellow Mustard [PYM] and Tillecur [Ti]) as well as milled Chinese galls (CG). At 2% concentration (wt/vol), PYM and Ti completely inhibited growth of mycelium of all F. graminearum strains whereas, at 1%, CG reduced the growth by 65 to 83%, depending on the strain. While PYM and Ti reduced the germination of both conidia and ascospores at 2% (wt/vol), CG was only effective in reducing conidia germination. Perithecia formation of FG0410 but not FG2113 was suppressed by all botanicals. Moreover, application of botanicals on mature perithecia led to a two- to fourfold reduction in discharge of ascospores. Using liquid chromatography (LC) with diode array detection, we quantified the principal glucosinolate component sinalbin of PYM and Ti. LC time-of-flight mass spectrometry was used to demonstrate that the bioactive matrix of CG contains different gallotannins as well as gallic and tannic acids. Possible antifungal mechanisms of the botanical matrices are discussed. The results of this study are promising and suggest that PYM, Ti, and CG should be explored further for efficacy at managing FHB.[Formula: see text] Copyright © 2019 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Development of a Bioactive Sauce Based on Oriental Mustard Flour with Antifungal Properties for Pita Bread Shelf Life Improvement.

Ochratoxin A (OTA) is a mycotoxin produced in the secondary metabolism of fungus belonging to the genus Aspergillus and Penicillium. In this study, the employment of oriental mustard flour (OMF) as an ingredient in a packaged sauce was evaluated for the generation in situ of the antimicrobial compound allyl isothiocyanate (AITC) in order to preserve pita bread contaminated with Penicillium verrucosum VTT D-01847, an OTA producer, in an active packaging system. Four different concentrations (8, 16, 33 and 50 mg/g) were tested. Mycelium formation, mycotoxin production, AITC absorbed by the food matrix, and volatilization kinetics were studied for each concentration. The results obtained were compared with bread treated with the commercial additive calcium propionate (E-282). The results showed a shelf life increase of two and three days with the employment of 33 and 50 mg/g of OMF, with a significant reduction of the fungal population (3.1 and 5.7 logs, respectively) in comparison with the control experiment. The use of 16 and 33 mg/g of OMF in the sauce formulation decreased the concentration of OTA in the bread samples while no OTA production was detected employing 50 mg/g of OMF.

Potential Application of Lactic Acid Bacteria to Reduce Aflatoxin B1 and Fumonisin B1 Occurrence on Corn Kernels and Corn Ears.

Fungal spoilage is an important issue for the food industry, leading to food sensory defects, food waste, economic losses and public health concern through the production of mycotoxins. Concomitantly, the search for safer natural products has gained importance since consumers began to look for less processed and chemically treated foods. In this context, the aim of this study was to evaluate the antifungal and antimycotoxigenic effect of seven strains of Lactobacillus plantarum. Lactic acid bacteria (LAB) were grown on Man Rogosa Sharpe (MRS) broth at 37 ºC in anaerobic conditions. After that, the cell-free supernatant (CFS) were recovered to determine its antifungal activity by halo diffusion agar test. In addition, minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) was determined for each L. plantarum CFS by 96-well microplates method. Additionally, CFS was used as a natural biocontrol agent on corn kernels and corn ears contaminated with Aspergillus flavus and Fusarium verticillioides, respectively. The L. plantarum CECT 749 CFS showed the highest antifungal effect against all essayed strains. Moreover, the employment of this CFS in food reduced the mycotoxin production at a percentage ranging from 73.7 to 99.7%. These results suggest that the L. plantarum CECT 749 CFS could be promising for the biocontrol of corn.

Aflatoxins and A. flavus Reduction in Loaf Bread through the Use of Natural Ingredients.

In this study, the antifungal activity of yellow mustard (YMF) and oriental mustard (OMF) meal extracts against 14 strains of fungi was tested on a solid medium. The results obtained with the YMF were next confirmed in liquid medium determining the minimum inhibitory concentration (MIC) and the minimum fungicide concentration (MFC). Finally, the use of YMF as a natural preservative to extend the useful life of bread was evaluated. Breads with different concentrations of YMF (2, 4, 6 and 8 g/kg) were prepared and contaminated with Aspergillus flavus ISPA 8111 and Penicilliumnordicum CECT 2320. For 10 days the formation of mycelium was observed, and after that the fungal growth and the mycotoxins production was determined. The results obtained with the YMF were compared with breads treated with the commercial additive sodium propionate (E-281). The results showed a significant reduction of the fungal population using 6 g/kg and 8 g/kg of YMF in bread contaminated with A. flavus and with P.nordicum and an extensions of the breads shelf life of 7 and 5 days, respectively, in comparison with the control experiment. A reduction of 78% of AFB1 was observed using 6 g/kg of YMF while no AFB1 production was detected employing 8 g/kg of YMF in bread preparation.