Bacillus velezensis CL197: a zearalenone detoxifying strain isolated from wheat with potential to be used in animal production.

Zearalenone (ZEA) is a mycotoxin produced by Fusarium species, and cause contamination of food and feed, with impacts in animal production and in food production chain. Effective detoxifying methods, such as biodegradation, are therefore required. This study aimed to isolate microorganisms and screen ZEA detoxifying strains. As a result, 197 microorganisms were isolated, and six were initially selected after colorimetric screening. ZEA (1 µg/mL) was added to culture media, and after 24 h, all six microorganisms were able to degrade ZEA, without the formation of α-ZOL. One isolate eliminated ~ 99% of ZEA and was identified as Bacillus velezensis CL197. ZEA metabolites produced by the bacteria were evaluated, and no metabolites with greater or similar toxicity than ZEA were detected. This strain was applied to swine in vitro digestion, and up to 64% of ZEA was degraded. B. velezensis CL197 significantly degraded ZEA, demonstrating potential to be used as a detoxifying agent in the food production chain as a biocontrol agent.

Simultaneous analysis of antimicrobial residues and contaminants in poultry droppings by HPLC-MS/MS: a tool for environmental and food safety monitoring.

Animal waste is a potential pollution hazard as it can harbour contaminants, such as antimicrobial residues, mycotoxins, and pesticides, becoming a risk to the public, animal, and environmental health. To assess this risk, 15 experimental broiler chickens orally received contaminants to evaluate excretion levels. An analytical method was previously developed to detect 18 substances in poultry droppings using high-performance liquid chromatography coupled to a tandem mass spectrometer (HPLC-MS/MS). Contaminants including tetracycline, 4-epi-tetracycline, oxytetracycline, 4-epi-oxytetracycline, chlortetracycline, 4-epi-chlortetracycline, tylosin, erythromycin, enrofloxacin, ciprofloxacin, flumequine, florfenicol, sulfachloropyridazine, sulfadiazine, 2,4-dichlorophenoxyacetic acid, zearalenone, alpha- and beta-zearalenol, were extracted with EDTA-McIlvain and acetonitrile. This method showed a p-value < 0.05, RSD < 25%, and R2 > 0.95 in the calibration curves linearity for all analytes. The limit of quantification, selectivity, decision limit for confirmation, matrix effect, precision, and recovery parameters were validated according to European Union document 2021/808/EC, technical report CEN/TR 16059, SANTE/11813/2017 and according to the Veterinary International Conference on Harmonization: VICH GL2 and GL49. This method confirmed the detection of most analytes 12-36 h post-administration and simultaneously detected and quantified mixed contaminants. Thereby, poultry droppings are a potential matrix for spreading contaminants in animal production before slaughter and their control will minimize environmental impacts and mitigate antimicrobial resistance.

Increased Dissemination of Aflatoxin- and Zearalenone-Producing Aspergillus spp. and Fusarium spp. during Wet Season via Houseflies on Dairy Farms in Aguascalientes, Mexico.

Crops contamination with aflatoxins (AFs) and zearalenone (ZEA) threaten human and animal health; these mycotoxins are produced by several species of Aspergillus and Fusarium. The objective was to evaluate under field conditions the influence of the wet season on the dissemination of AF- and ZEA-producing fungi via houseflies collected from dairy farms. Ten dairy farms distributed in the semi-arid Central Mexican Plateau were selected. Flies were collected in wet and dry seasons at seven points on each farm using entomological traps. Fungi were isolated from fly carcasses via direct seeding with serial dilutions and wet chamber methods. The production of AFs and ZEA from pure isolates was quantified using indirect competitive ELISA. A total of 693 Aspergillus spp. and 1274 Fusarium spp. isolates were obtained, of which 58.6% produced AFs and 50.0% produced ZEA (491 ± 122; 2521 ± 1295 µg/kg). Houseflies and both fungal genera were invariably present, but compared to the dry season, there was a higher abundance of flies as well as AF- and ZEA-producing fungi in the wet season (p < 0.001; 45.3/231 flies/trap; 8.6/29.6% contaminated flies). These results suggest that rainy-weather conditions on dairy farms increase the spread of AF- and ZEA-producing Aspergillus spp. and Fusarium spp. through houseflies and the incorporation of their mycotoxins into the food chain.

Occurrence of ochratoxin A in cocoa beans and bean-to-bar chocolates.

The growing health consciousness of consumers has led to an increase in the consumption of artisanal chocolates, mainly due to their recognized health benefits. However, processing steps such as fermentation and drying of cocoa beans can favor the growth of ochratoxigenic fungi. This study aimed to assess the occurrence of ochratoxin A (OTA) in cocoa beans (purchased from e-commerce and post-harvest processing) and bean-to-bar chocolates sold in Brazil. An HPLC-FLD method was validated, with recovery values between 84 and 97% and limits of detection and quantification of 0.04 and 0.01 µg/kg, respectively. OTA was detected in 30% of the cocoa bean samples studied (n = 43), with values ranging from < 0.04 to 1.18 µg/kg. Regarding the bean-to-bar chocolates (n = 62), the OTA concentrations ranged from < 0.04 to 1.11 µg/kg, with a prevalence in semi-sweet and dark chocolates. Despite representing a growing market, to the best of our knowledge, this is the first study to report OTA concentrations in bean-to-bar chocolates and Brazilian cocoa beans used to produce this type of chocolate.

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.

Essential oils: An eco-friendly alternative for controlling toxigenic fungi in cereal grains.

Fungi are widely disseminated in the environment and are major food contaminants, colonizing plant tissues throughout the production chain, from preharvest to postharvest, causing diseases. As a result, grain development and seed germination are affected, reducing grain quality and nutritional value. Some fungal species can also produce mycotoxins, toxic secondary metabolites for vertebrate animals. Natural compounds, such as essential oils, have been used to control fungal diseases in cereal grains due to their antimicrobial activity that may inhibit fungal growth. These compounds have been associated with reduced mycotoxin contamination, primarily related to reducing toxin production by toxigenic fungi. However, little is known about the mechanisms of action of these compounds against mycotoxigenic fungi. In this review, we address important information on the mechanisms of action of essential oils and their antifungal and antimycotoxigenic properties, recent technological strategies for food industry applications, and the potential toxicity of essential oils.

Nursery pigs fed with feed contaminated by aflatoxin B1 (Aspergillus flavus) and anti-mycotoxin blend: Pathogenesis and negative impact on animal health and weight gain.

The present study aimed to evaluate whether a moderate dose of aflatoxin B1 in pigs' diet interferes with pigs' growth and health in the nursery phase and whether an anti-mycotoxin mixture minimizes the adverse effects of the toxin. One blend with Saccharomyces cerevisiae lysate, zeolite, silicon dioxide, propylene glycol, Carduus marianus extract, soy lecithin, and carbonate was used as an anti-mycotoxin. Four treatments, with six repetitions per treatment and three pigs/pen: Afla0-AntiMyc0 - negative control (without aflatoxin); Afla500-AntiMyc0 - positive control (500 ppb of aflatoxin); Afla0-AntiMyc1000 - 1000 mg/kg of anti-mycotoxin blend; Afla500-AntiMyc1000 - 500 ppb aflatoxin +1000 mg/kg of anti-mycotoxin blend. It was observed that pigs in the positive control (Afla500-AntiMyc0) had lower body weight and weight gain when compared to the other treatments during the experimental period. Also, pigs from Afla500-AntiMyc0 had lower feed intake between days 1-20 and 1 to 30 than Afla0-AntiMyc0. The pigs from Afla500-AntiMyc0 had higher levels of liver enzymes aspartate aminotransferase and alanine aminotransferase compared to other treatments. The pigs from Afla500-AntiMyc0 had higher villus height than the other treatments, while the folded size was smaller in this treatment. Crypts were deeper in the intestines of pigs in both treatments that consumed aflatoxin. In general, it is concluded that the intake of aflatoxin B1 by nursery pigs has negative impacts on the health and, consequently, the animals' growth performance; however, the addition of the contaminated feed with an anti-mycotoxin blend was able to protect the pigs, minimizing the adverse effects caused by the mycotoxin.

Investigation of the mutagenic, cytotoxic and antimicrobial effect of bikaverin mycotoxin

Bikaverin is a reddish pigment produced by different fungi species (Mycogone jaapii, Verticillium agaricinum, Beauveria bassiana, Paecilomyces fumosoroseus, Polyporus sulphureus), mainly of the Fusarium genus. Due to its pigment feature, bikaverin can be used as a dye in various fields in the industry. However, it is extremely important to study the mutagenic/genotoxic effects, cytotoxic effects and antimicrobial properties of bikaverin for application of industrial areas. In the study, the mutagenic, cytotoxic and antimicrobial effects of bikaverin were investigated. The mutagenic effect of bikaverin was studied with the Ames test. Salmonella typhimurium TA97a, TA98, TA100, TA102 and TA1535 strains were used in the test. Five different doses of bikaverin (0.075, 0.1, 0.2, 0.3 and 0.5 µg/plate) were tested against strains. It was determined that there was no mutagenic effect of bikaverin. The cytotoxicity of bikaverin was evaluated by MTT test on L929 fibroblast cell line. Bikaverin demonstrated no cytotoxic effect on L929 fibroblast cell line, according to cell viability calculations that showed >73% for all concentrations (1, 0.5, 0.4, 0.3, 0.2, 0.1, 0.075, 0.05, 0.025, 0.01, 0.005 and 0.001 µg/mL) examined. Bikaverin's IC50 value was determined to be 1.79±0.51 g/mL. The antimicrobial activity of the bikaverin was evaluated by using the microdilution method. Bikaverin was found to have antimicrobial effects on Methicillin resistant Staphylococcus aureus, Vancomycin resistant Enterococcus faecalis, Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, Candida albicans and Candida krusei, as MIC values ranged from 1.25 -5 µg/ mL.

Assessment of quality and safety aspects of homemade and commercial baby foods.

Dietary Guidelines in some countries recommend avoiding commercially processed baby food, while others encourage the consultation of ingredients and nutritional information. Therefore, the objective of this study was to systematically analyze different baby foods obtained from commercial market and "homemade" produced, in order to verify whether comercial products have low nutritional and unsafety attributes. The samples were analyzed for chemical composition, physicochemical aspects, texture, microbiological and mycotoxin contamination, and pesticide residues. Results showed that, in general, commercial samples have a higher energy density and better ratio of macronutrients. The sodium, pH, and texture of both products were in accordance with the recommendations. None of the baby foods evaluated were contaminated with yeast and molds, total coliforms, or Escherichia coli; however, Salmonella sp. was confirmed in one homemade sample. Pesticide residues were detected in all analyzed baby food samples; however, at lower levels than the limit of quantification. Ochratoxin A was detected in one homemade baby food sample (5.76 µg /kg). Considering the samples evaluated, commercial baby food samples appeared to be safer in relation to microbiological, pesticide residue standards, and mycotoxin contamination. Therefore, it could be concluded that the quality of commercial and homemade baby foods still needs to be improved, as well as more studies related to a critical analyses of both types of processes used.

Evaluation of Interactions of Added Soybean Peroxidase with Other Nutrients Present in Fish Feeds Using an In Vitro Digestive Simulation.

Peroxidase (PO) has been applied in different areas of industrial biotechnology, including the control of contaminants like aflatoxin B1 in fish feeds. However, its potential negative interactions with the macro and micro components of feeds have not been evaluated. The aim of this study was to evaluate the impact of PO's addition to a feed on compounds like fatty acids and polyphenols using an in vitro simulation of the digestive tract of the tilapia. The influence on fatty acids was determined by changes in the peroxide index, with the feed including PO presenting values four times higher than those of the control feed. On the other hand, the in vitro digestive simulation also evidenced an effect of PO on the bioaccessibility of polyphenols significantly influenced by the total digestion time and temperature. The bioaccessibility of polyphenol ranged from 2.09 to 16.23 µmol of the total Trolox equivalent antioxidant capacity for the combinations evaluated in the study. The greatest bioaccessibility was observed at the central point under the following conditions of digestive hydrolysis: pH of 7, 30 °C, 4.5 h of digestive hydrolysis and an absence of PO.

Genetic Diversity of Aspergillus flavus on Maize in Guatemala.

Aflatoxin contamination of maize is a leading threat to health in Guatemala. This contamination is the result of infection from Aspergillus flavus and has been effectively reduced in other countries through application of nonaflatoxigenic, indigenous strains of A. flavus. We collected 82 maize samples from throughout Guatemala in two years and isolated 272 A. flavus from these samples, including 126 unique genotypes. We provide here a phenotypic and simple sequence repeat (SSR)-based genotypic description of these isolates, as well as an analysis of the diversity of this population. High levels of genetic diversity were observed with the nonaflatoxigenic isolates in this study, but this information contributes to the development of indigenous aflatoxin biocontrol products.

Pitaya Juice Consumption Protects against Oxidative Damage Induced by Aflatoxin B1.

Mycotoxins are toxic fungal metabolites and are responsible for contaminating several foods. The intake of foods contaminated by these substances is related to hepatotoxicity and carcinogenic effects, possibly due to increasing oxidative stress. The current study evaluated Pitaya fruit juice's antioxidant effects on oxidative damage aflatoxin B1 (AFB1)-induced. Rats received 1.5 mL of Pitaya juice via gavage (for 30 days), and on the 31st day, they received AFB1 (250 µg/kg, via gavage). Forty-eight hours after the AFB1 dose, rats were euthanized for dosages of alanine transaminase (ALT), aspartate aminotransferase (AST), and alkaline phosphatase (ALP); dosage of oxidative markers (thiobarbituric acid reactive species (TBARS), reactive species (RS)) and antioxidant defenses (catalase (CAT), superoxide dismutase (SOD), Glutathione S-transferase (GST) activities and Glutathione (GSH)) levels in the liver; and detection of Heat shock protein 70 (Hsp-70) and nuclear factor- erythroid 2-related factor 2 (Nrf2) immunocontent in the liver. Our results indicated that the Pitaya juice reduced ALP activity. Further, rats exposed to AFB1 experienced liver damage due to the increase in TBARS, RS, and Hsp-70 and the reduction in CAT, GSH, and Nrf2. Pitaya juice could, however, protect against these damages. Finally, these results indicated that pre-treatment with Pitaya juice was effective against the oxidative damage induced. However, other aspects may be elucidated in the future to discover more targets of its action against mycotoxicosis.

Yeasts as sustainable biocontrol agents against ochratoxigenic Aspergillus species and in vitro optimization of ochratoxin A detoxification.

AIMS: The aims of this study were to evaluate the potential of Hanseniaspora opuntiae, Meyerozyma caribbica, and Kluyveromyces marxianus for in vitro biocontrol of Aspergillus ochraceus, A. westerdijkiae, and A. carbonarius growth, the ochratoxin A (OTA) effect on yeast growth, and yeast in vitro OTA detoxification ability using an experimental design to predict the combined effects of inoculum size, incubation time, and OTA concentration. METHODS AND RESULTS: Predictive models were developed using an incomplete Box-Behnken experimental design to predict the combined effects of inoculum size, incubation time, and OTA concentration on OTA detoxification by the yeasts. The yeasts were able to inhibit fungal growth from 13% to 86%. Kluyveromyces marxianus was the most efficient in inhibiting the three Aspergillus species. Furthermore, high OTA levels (100 ng ml-1) did not affect yeast growth over 72 h incubation. The models showed that the maximum OTA detoxification under optimum conditions was 86.8% (H. opuntiae), 79.3% (M. caribbica), and 73.7% (K. marxianus), with no significant difference (P > 0.05) between the values predicted and the results obtained experimentally. CONCLUSION: The yeasts showed potential for biocontrol of ochratoxigenic fungi and OTA detoxification, and the models developed are important tools for predicting the best conditions for the application of these yeasts as detoxification agents.

Species Identification and Mycotoxigenic Potential of Aspergillus Section Flavi Isolated from Maize Marketed in the Metropolitan Region of Asunción, Paraguay.

Zea mays var. amylacea and Zea mays var. indurata are maize ecotypes from Paraguay. Aspergillus section Flavi is the main spoilage fungus of maize under storage conditions. Due to its large intraspecific genetic variability, the accurate identification of this fungal taxonomic group is difficult. In the present study, potential mycotoxigenic strains of Aspergillus section Flavi isolated from Z. mays var. indurata and Z. mays var. amylacea that are marketed in the metropolitan region of Asunción were identified by a polyphasic approach. Based on morphological characters, 211 isolates were confirmed to belong to Aspergillus section Flavi. A subset of 92 strains was identified as Aspergillus flavus by mass spectrometry MALDI-TOF and the strains were classified by MALDI-TOF MS into chemotypes based on their aflatoxins and cyclopiazonic acid production. According to the partial sequencing of ITS and CaM genes, a representative subset of 38 A. flavus strains was confirmed. Overall, 75 A. flavus strains (86%) were characterized as producers of aflatoxins. The co-occurrence of at least two mycotoxins (AF/ZEA, FUM/ZEA, and AF/ZEA/FUM) was detected for five of the Z. mays samples (63%). Considering the high mycological bioburden and mycotoxin contamination, maize marketed in the metropolitan region of Asunción constitutes a potential risk to food safety and public health and requires control measures.

Study of co-occurrence of mycotoxins in cocoa beans in Brazil by liquid chromatography-tandem mass spectrometry (LC-MS/MS).

In this study, 135 samples of cocoa beans collected in the Amazon and Atlantic Forest regions of Brazil were analysed to evaluate the possible co-occurrence of 34 mycotoxins. The results indicate that 42% of the cocoa samples exhibited quantifiable levels for 11 mycotoxins: aflatoxins (AFs) B1, B2 and G1; ochratoxin A; citrinin; cyclopiazonic acid; tenuazonic acid; paxilline; sterigmatocystin; zearalenone and fumonisin B2. Of the samples, 18% exhibited the co-occurrence of up to six mycotoxins. No toxins belonging to the groups of trichothecenes or ergot alkaloids were detected. Contingency analysis of the incidence of mycotoxins did not show significant differences between the two regions evaluated. Seven samples were contaminated with AFs, while only one contained ochratoxin A above 10 µg kg-1. The accuracy of the method was evaluated by proficiency testing for ochratoxin A, where satisfactory Z-scores were obtained.

Molecular Docking and In Vitro Studies of Ochratoxin A (OTA) Biodetoxification Testing Three Endopeptidases.

Ochratoxin A (OTA) is considered one of the main mycotoxins responsible for health problems and considerable economic losses in the feed industry. The aim was to study OTA's detoxifying potential of commercial protease enzymes: (i) Ananas comosus bromelain cysteine-protease, (ii) bovine trypsin serine-protease and (iii) Bacillus subtilis neutral metalloendopeptidase. In silico studies were performed with reference ligands and T-2 toxin as control, and in vitro experiments. In silico study results showed that tested toxins interacted near the catalytic triad, similar to how the reference ligands behave in all tested proteases. Likewise, based on the proximity of the amino acids in the most stable poses, the chemical reaction mechanisms for the transformation of OTA were proposed. In vitro experiments showed that while bromelain reduced OTA's concentration in 7.64% at pH 4.6; trypsin at 10.69% and the neutral metalloendopeptidase in 8.2%, 14.44%, 45.26% at pH 4.6, 5 and 7, respectively (p < 0.05). The less harmful α-ochratoxin was confirmed with trypsin and the metalloendopeptidase. This study is the first attempt to demonstrate that: (i) bromelain and trypsin can hydrolyse OTA in acidic pH conditions with low efficiency and (ii) the metalloendopeptidase was an effective OTA bio-detoxifier. This study confirmed α-ochratoxin as a final product of the enzymatic reactions in real-time practical information on OTA degradation rate, since in vitro experiments simulated the time that food spends in poultry intestines, as well as their natural pH and temperature conditions.

Zeolite and corn with different compositions in broiler chickens feeding.

The objective of this work was to evaluate the behavior of zeolite against different types of corn in broiler chickens' diets. 1,200 male broiler chickens were assigned to 6 diets in a completely randomized design and a 2 × 3 factorial arrangement, consisting of 2 types of corn (higher or lower mycotoxin contamination) and the inclusion of zeolite (0; 5,000 and 10,000 g ton-1). In the period from d 1 to 21, there was an interaction (P = 0.0040) between types of corn and the inclusion of zeolite for feed conversion ratio (FCR). In the phase from 1 to 42 d, there was an interaction (P = 0.0322) on the serum levels of creatinine (CREA) and digestible gross energy (dGE); corn with lower mycotoxin level contamination (LMLC), caused a reduction in body weight gain (BWG) (P = 0.0046) and increase in the relative weight of abdominal fat (P = 0.0256). Inclusion rates of zeolite promoted an increase in the digestible CP (P = 0.0477) and digestible ash (P < 0.0001), as well as an increase (P < 0.0001) in hot carcass yield (P = 0.0433). The results indicate that the inclusion of zeolite, in the amounts used, did not alter the performance, serum levels, intestinal development and litter quality of the birds at 42 d of age. However, it was responsible for the improvement in the percentage of digestible nutrients (CP, GE, and MM).

Computational Studies of Aflatoxin B1 (AFB1): A Review.

Aflatoxin B1 (AFB1) exhibits the most potent mutagenic and carcinogenic activity among aflatoxins. For this reason, AFB1 is recognized as a human group 1 carcinogen by the International Agency of Research on Cancer. Consequently, it is essential to determine its properties and behavior in different chemical systems. The chemical properties of AFB1 can be explored using computational chemistry, which has been employed complementarily to experimental investigations. The present review includes in silico studies (semiempirical, Hartree-Fock, DFT, molecular docking, and molecular dynamics) conducted from the first computational study in 1974 to the present (2022). This work was performed, considering the following groups: (a) molecular properties of AFB1 (structural, energy, solvent effects, ground and the excited state, atomic charges, among others); (b) theoretical investigations of AFB1 (degradation, quantification, reactivity, among others); (c) molecular interactions with inorganic compounds (Ag+, Zn2+, and Mg2+); (d) molecular interactions with environmentally compounds (clays); and (e) molecular interactions with biological compounds (DNA, enzymes, cyclodextrins, glucans, among others). Accordingly, in this work, we provide to the stakeholder the knowledge of toxicity of types of AFB1-derivatives, the structure-activity relationships manifested by the bonds between AFB1 and DNA or proteins, and the types of strategies that have been employed to quantify, detect, and eliminate the AFB1 molecule.

Mixtures of Mycotoxins, Phytoestrogens, and Other Secondary Metabolites in Whole-Plant Corn Silages and Total Mixed Rations of Dairy Farms in Central and Northern Mexico.

Mycotoxins and endocrine disruptors such as phytoestrogens can affect cattle health, reproduction, and productivity. Most studies of mycotoxins in dairy feeds in Mexico and worldwide have been focused on a few (regulated) mycotoxins. In contrast, less known fungal toxins, phytoestrogens, and other metabolites have been neglected and underestimated. This study analyzed a broad spectrum (>800) of mycotoxins, phytoestrogens, and fungal, plant, and unspecific secondary metabolites in whole-plant corn silages (WPCSs) and total mixed rations (TMRs) collected from 19 Mexican dairy farms. A validated multi-metabolite liquid chromatography/electrospray ionization-tandem mass spectrometric (LC/ESI-MS/MS) method was used. Our results revealed 125 of >800 tested (potentially toxic) secondary metabolites. WPCSs/TMRs in Mexico presented ubiquitous contamination with mycotoxins, phytoestrogens, and other metabolites. The average number of mycotoxins per TMR was 24, ranging from 9 to 31. Fusarium-derived secondary metabolites showed the highest frequencies, concentrations, and diversity among the detected fungal compounds. The most frequently detected mycotoxins in TMRs were zearalenone (ZEN) (100%), fumonisin B1 (FB1) (84%), and deoxynivalenol (84%). Aflatoxin B1 (AFB1) and ochratoxin A (OTA), previously reported in Mexico, were not detected. All TMR samples tested positive for phytoestrogens. Among the investigated dietary ingredients, corn stover, sorghum silage, and concentrate proportions were the most correlated with levels of total mycotoxins, fumonisins (Fs), and ergot alkaloids, respectively.

Repeated co-exposure to aflatoxin B1 and aspartame disrupts the central nervous system homeostasis: Behavioral, biochemical, and molecular insights.

Several studies demonstrated the toxicity of aspartame (ASP) and aflatoxin B1 (AFB1 ) in preclinical models. Although the majority of these reports assessed the toxic effects of each substance separately, their concomitant exposure and hazardous consequences are scarce. Importantly, the deleterious effects at the central nervous system caused by ASP and AFB1 co-exposure are rarely addressed. We evaluated if concomitant exposure to AFB1 and ASP would cause behavioral impairment and alteration in oxidative status of the brain in male rats. Animals received once a day for 14 days AFB1 (250 µg/kg, intragastric gavage [i.g.]), ASP (75 mg/kg, i.g.), or both substances (association). On day 14, they were subjected to behavioral evaluation, and biochemical and molecular parameters of oxidative status were measured in the cerebral cortex and hippocampus. In the open field test, AFB1 and combination treatments modified the motor, exploratory, and grooming behavior. In the splash test, all treatments caused a reduction in grooming time compared to the control group. An increase in thiobarbituric acid-reactive substances content induced by AFB1 and combination treatments was observed. The antioxidant defenses (vitamin C, nonprotein sulfhydryl, and ferric reducing antioxidant power) were impaired in all groups compared to control. Regarding molecular evaluation, mitochondrial superoxide dismutase-2 immunoreactivity decreased after AFB1 or ASP exposition in the hippocampus. Thus, co-exposure to ASP and AFB1 was potentially more toxic because it aggravated behavioral impairments and oxidative status disbalance in comparison to the groups that received only ASP or AFB1 . Therefore, our data suggest that those substances caused a disruption in brain homeostasis.