Mycotoxins are secondary metabolites produced by fungi especially those belonging to the genus Aspergillus, Penicillum and Fusarium. Mycotoxin contamination can occur in all agricultural commodities in the field and/or during storage, if conditions are favourable to fungal growth. Regarding animal feed, five mycotoxins (aflatoxins, deoxynivalenol, zearalenone, fumonisins and ochratoxin A) are covered by EU legislation (regulation or recommendation). Transgressions of these limits are rarely observed in official monitoring programs. However, low level contamination by Fusarium toxins is very common (e.g., deoxynivalenol (DON) is typically found in more than 50% of the samples) and co-contamination is frequently observed. Multi-mycotoxin studies reported 75%–100% of the samples to contain more than one mycotoxin which could impact animal health at already low doses. Co-occurrence of mycotoxins is likely to arise for at least three different reasons (i) most fungi are able to simultaneously produce a number of mycotoxins, (ii) commodities can be contaminated by several fungi, and (iii) completed feed is made from various commodities. In the present paper, we reviewed the data published since 2004 concerning the contamination of animal feed with single or combinations of mycotoxins and highlighted the occurrence of these co-contaminations.

Patulin is a toxic chemical contaminant produced by several species of mold, especially within Aspergillus, Penicillium and Byssochlamys. It is the most common mycotoxin found in apples and apple-derived products such as juice, cider, compotes and other food intended for young children. Exposure to this mycotoxin is associated with immunological, neurological and gastrointestinal outcomes. Assessment of the health risks due to patulin consumption by humans has led many countries to regulate the quantity in food. A full understanding of the molecular genetics of patulin biosynthesis is incomplete, unlike other regulated mycotoxins (aflatoxins, trichothecenes and fumonisins), although the chemical structures of patulin precursors are now known. The biosynthetic pathway consists of approximately 10 steps, as suggested by biochemical studies. Recently, a cluster of 15 genes involved in patulin biosynthesis was reported, containing characterized enzymes, a regulation factor and transporter genes. This review includes information on the current understanding of the mechanisms of patulin toxinogenesis and summarizes its toxicological effects.

The study of fungal species evolved radically with the development of molecular techniques and produced new evidence to understand specific fungal mechanisms such as the production of toxic secondary metabolites. Taking advantage of these technologies to improve food safety, the molecular study of toxinogenic species can help elucidate the mechanisms underlying toxin production and enable the development of new effective strategies to control fungal toxicity. Numerous studies have been made on genes involved in aflatoxin B1 (AFB1) production, one of the most hazardous carcinogenic toxins for humans and animals. The current review presents the roles of these different genes and their possible impact on AFB1 production. We focus on the toxinogenic strains Aspergillus flavus and A. parasiticus, primary contaminants and major producers of AFB1 in crops. However, genetic reports on A. nidulans are also included because of the capacity of this fungus to produce sterigmatocystin, the penultimate stable metabolite during AFB1 production. The aim of this review is to provide a general overview of the AFB1 enzymatic biosynthesis pathway and its link with the genes belonging to the AFB1 cluster. It also aims to illustrate the role of global environmental factors on aflatoxin production and the recent data that demonstrate an interconnection between genes regulated by these environmental signals and aflatoxin biosynthetic pathway.

Mycotoxins are secondary fungal metabolites produced mainly by Aspergillus, Penicillium, and Fusarium. As evidenced by large-scale surveys, humans and animals are simultaneously exposed to several mycotoxins. Simultaneous exposure could result in synergistic, additive or antagonistic effects. However, most toxicity studies addressed the effects of mycotoxins separately. We present the experimental designs and we discuss the conclusions drawn from in vitro experiments exploring toxicological interactions of mycotoxins. We report more than 80 publications related to mycotoxin interactions. The studies explored combinations involving the regulated groups of mycotoxins, especially aflatoxins, ochratoxins, fumonisins, zearalenone and trichothecenes, but also the "emerging" mycotoxins beauvericin and enniatins. Over 50 publications are based on the arithmetic model of additivity. Few studies used the factorial designs or the theoretical biology-based models of additivity. The latter approaches are gaining increased attention. These analyses allow determination of the type of interaction and, optionally, its magnitude. The type of interaction reported for mycotoxin combinations depended on several factors, in particular cell models and the tested dose ranges. However, synergy among Fusarium toxins was highlighted in several studies. This review indicates that well-addressed in vitro studies remain valuable tools for the screening of interactive potential in mycotoxin mixtures.

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Deoxynivalenol (DON) is the most prevalent trichothecene mycotoxin in crops in Europe and North America. DON is often present with other type B trichothecenes such as 3-acetyldeoxynivalenol (3-ADON), 15-acetyldeoxynivalenol (15-ADON), nivalenol (NIV) and fusarenon-X (FX). Although the cytotoxicity of individual mycotoxins has been widely studied, data on the toxicity of mycotoxin mixtures are limited. The aim of this study was to assess interactions caused by co-exposure to Type B trichothecenes on intestinal epithelial cells. Proliferating Caco-2 cells were exposed to increasing doses of Type B trichothecenes, alone or in binary or ternary mixtures. The MTT test and neutral red uptake, respectively linked to mitochondrial and lysosomal functions, were used to measure intestinal epithelial cytotoxicity. The five tested mycotoxins had a dose-dependent effect on proliferating enterocytes and could be classified in increasing order of toxicity: 3-ADON<15-ADON≈DON

Inhalation of Aspergillus fumigatus conidia can cause severe aspergillosis in immunosuppressed people. A. fumigatus produces a large number of secondary metabolites, some of which are airborne by conidia and whose toxicity to the respiratory tract has not been investigated. We found that spores of A. fumigatus contain five main compounds, tryptoquivaline F, fumiquinazoline C, questin, monomethylsulochrin and trypacidin. Fractionation of culture extracts using RP-HPLC and LC-MS showed that samples containing questin, monomethylsulochrin and trypacidin were toxic to the human A549 lung cell line. These compounds were purified and their structure verified using NMR in order to compare their toxicity against A549 cells. Trypacidin was the most toxic, decreasing cell viability and triggering cell lysis, both effects occurring at an IC50 close to 7 µM. Trypacidin toxicity was also observed in the same concentration range on human bronchial epithelial cells. In the first hour of exposure, trypacidin initiates the intracellular formation of nitric oxide (NO) and hydrogen peroxide (H2O2). This oxidative stress triggers necrotic cell death in the following 24 h. The apoptosis pathway, moreover, was not involved in the cell death process as trypacidin did not induce apoptotic bodies or a decrease in mitochondrial membrane potential. This is the first time that the toxicity of trypacidin to lung cells has been reported.

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