Mycotoxines

The impact of chlorophyllin on deoxynivalenol transport across jejunum mucosa explants obtained from adult pigs Abstract Regardless of the efforts put into preventing or reducing fungal growth, extensive mycotoxin contamination has been reported in animal feeds. In the case of pigs, one of the mycotoxins of major concern is deoxynivalenol (DON). The use of adsorbents as feed additives represents one of the strategies to control mycotoxins' contamination in feedstuff. Therefore, the aim of the study was to verify the ability of chlorophyllin (CHL) to reduce the absorption rate of DON in swine mucosa explants. Intestine was obtained from routinely slaughtered adult pigs. The mucosa explants were studied by means of Ussing chamber technique. The effect of DON (10 and 30 μg/ml) on mucosa viability and permeability and CHL (100 μg/ml) impact on DON (30 μg/ml) absorption was verified. The results revealed that mucosa explants isolated from adult animals remained unaffected for 90 min in the presence of DON in the lower concentration (10 μg/ml). Mycotoxin in the higher dose (30 μg/ml) increased mucosa permeability (decreased transepithelial electrical resistance value) and enhanced paracellular transport of lucifer yellow and mannitol but did not affect lactate dehydrogenase leakage. The introduction of CHL neither diminished the absorption rate of DON across swine mucosa explants nor prevented the toxic effects of DON on intestine. In conclusion, the results confirm the negative effect of DON on pig jejunum mucosa. However, the toxic effect of DON was observed only when it was used in relatively high doses. A promising adsorbent agent, CHL, failed to reduce the intensity of DON transport across intestine under in vitro conditions.

Frequency and levels of mycotoxins in beer from the Mexican market and exposure estimate for deoxynivalenol mycotoxins Abstract The aim of the present study was to evaluate the occurrence of 23 mycotoxins in beer purchased in Mexico and to assess two exposure scenarios in the Mexican population through beer consumption. Multi-mycotoxin analysis of a total of 61 different beers (132 samples) was carried out using UHPLC-MS/MS equipment. Probability density functions were used to describe mycotoxins contamination. The daily intake of mycotoxins was estimated using a semi-probabilistic approach, applying the Monte Carlo method. Deoxynivalenol (DON) and its metabolites (deoxynivalenol-3-glucoside (DON3G) and 3-acetyl-deoxynivalenol (3ADON)) were the mycotoxins found in higher proportions in contaminated samples. None of the other mycotoxins overpassed the limit of quantification (LOQ) of the method. The combined intake of DON and its analogues ranged from 5.24 to 86.59 ng kg−1 bw day−1, which represent from 1.20 to 19.83% of the DON TDI. The results suggest that depending on the individual consumption of beer and depending on the type of beer, the intake of DON via beer could represent a significant percentage of the tolerable daily intake (TDI).

In vitro characterization of hepatic toxicity of Alternaria toxins Abstract Alternaria mycotoxins are secondary fungal metabolites which can contaminate food and feed. They are produced by Alternaria species with alternariol (AOH), alternariol monomethyl ether (AME), tenuazonic acid (TeA), and tentoxin (TEN) as the main representatives for Alternaria mycotoxins in food. Once passing the intestinal barrier, Alternaria toxins can reach the liver to exert yet uncharacterized molecular effects. Therefore, hepatic in vitro systems were used to examine selected Alternaria mycotoxins for their induction of metabolism-dependent cytotoxicity, phosphorylation of the histone H2AX as a surrogate marker for DNA double-strand breaks, and relevant marker genes for hepatotoxicity. Analysis of cell viability as well as the induction of H2AX phosphorylation in the hepatocarcinoma cell line HepG2 revealed a detoxification of 100 μmol/l AME and AOH by pre-treatment with S9 liver homogenate as shown by a decrease in cytotoxicity and H2AX histone phosphorylation to levels observed in control cells. Concentrations up to 100 μmol/l TeA and TEN did not induce H2AX phosphorylation whether metabolized or not. In the metabolically competent human hepatoma cell line HepaRG, no cytotoxicity of Alternaria toxins occurred even at high concentrations up to 100 μmol/l, which indicates a low cytotoxic potential. Induction of gene expression associated with liver toxicity was analyzed by quantitative real-time PCR using a specific hepatotoxicity PCR array in HepaRG cells: here, an evidence was found that 50 μmol/l of AOH, AME, TeA, and TEN might be associated with hepatotoxic effects, necrosis, and the development of diseases like cholestasis and phospholipidosis.

Occurrence of ochratoxin A in typical salami produced in different regions of Italy Abstract A total of 172 different salamis were purchased from farms and small salami factories located in four Italian regions (Piedmont, Veneto, Calabria, and Sicily) and analyzed for the presence of ochratoxin A (OTA). Analysis was performed by high-performance liquid chromatography coupled to a fluorimetric detector (HPLC-FLD). The detection limit (LOD) for the method used was 0.05 μg/kg, while the quantitation limit (LOQ) was 0.20 μg/kg; the average recovery rate was 89.1%. OTA was detected in 22 salamis, and 3 samples exceeded the Italian guidance value for OTA in pork meat (1 μg/kg). In particular, what emerges from this research is the high percentage of spicy salamis among positive samples (68.2%, 15 out of 22), although spicy salamis are only 27.3% of the total number of samples collected and analyzed. Red chili pepper contaminated by OTA could be responsible for the presence of the mycotoxin in these spicy salamis. It follow that, also the control of some ingredients used in the manufacture of these meat products, like spices, should not be neglected.

Transgenic versus conventional corn: fate of fumonisins during industrial dry milling Abstract The aim of this study was to compare the fate of fumonisins in transgenic and non-transgenic corn during industrial dry milling. For this purpose, whole corn samples and their fractions (germ, pericarp, endosperm, corn meal, and grits) were collected from one of the major Brazilian milling plants, totaling 480 samples. There was no significant difference (p > 0.05) between mean fumonisin (FB1 + FB2) levels in transgenic (1130 μg/kg) and non-transgenic (920 μg/kg) whole corn. However, in non-transgenic germ, endosperm and corn meal fraction fumonisin levels were higher (2940 μg/kg, 250 μg/kg and 190 μg/kg, respectively) than in transgenic fractions (2180 μg/kg, 130 μg/kg and 85.0 μg/kg, respectively). Furthermore, the highest percentages of fumonisins were distributed in the germ, corresponding to about 87 and 76% of the total fumonisins present in the whole corn from non-transgenic and transgenic hybrids, respectively. Concerning the endosperm from non-transgenic and transgenic corn, approximately, 23% and 13% of the total fumonisins were retained after the dry milling. Further processing in corn meal (300 to 420 μm particle size) and grits (590 to 1190 μm) decreased the percentages of remaining fumonisins to 4% and 2% (transgenic) and 10% and 3% (non-transgenic corn), respectively. These results suggested that fumonisin concentration was higher in outer and inner non-transgenic fractions when compared to transgenic ones and that the fate of fumonisins during the industrial dry milling could be affected by the transgenic status. However, it was not possible to conclude that the difference was exclusively due to this variable.

Interaction of the mycotoxin metabolite dihydrocitrinone with serum albumin Abstract Citrinin (CIT) is a nephrotoxic mycotoxin produced by Penicillium, Monascus, and Aspergillus species. CIT appears as a contaminant in cereals, cereal-based products, fruits, nuts, and spices. During the biotransformation of CIT, its major urinary metabolite dihydrocitrinone (DHC) is formed. Albumin interacts with several compounds (including mycotoxins) affecting their tissue distribution and elimination. CIT-albumin interaction is known; however, the complex formation of DHC with albumin has not been reported previously. In this study, we aimed to investigate the interaction of DHC with albumin, employing fluorescence spectroscopy, circular dichroism, and molecular modeling studies. Furthermore, species differences and thermodynamics of the interaction as well as the effects of albumin on the acute in vitro toxicity of DHC and CIT were also tested. Our main observations/conclusions are as follows: (1) Fluorescence signal of DHC is strongly enhanced by albumin. (2) Formation of DHC-albumin complexes is supported by both fluorescence spectroscopic and circular dichroism studies. (3) DHC forms similarly stable complexes with human albumin (K~105 L/mol) as CIT. (4) DHC-albumin interaction did not show significant species differences (tested with human, bovine, porcine, and rat albumins). (5) Based on modeling studies and investigations with site markers, DHC occupies the Heme binding site (subdomain IB) on human albumin. (6) The presence of albumin significantly decreased the acute in vitro cytotoxic effects of both DHC and CIT on MDCK cell line.

Aflatoxin in maize: a review of the early literature from "moldy-corn toxicosis" to the genetics of aflatoxin accumulation resistance Abstract Aflatoxin is a potent toxin produced by Aspergillus flavus Link:Fr, an opportunistic ear-rot pathogen of maize (Zea mays L. subsp. Mays). Prior to the discovery of aflatoxin, A. flavus was considered a minor pathogen and was not a priority for maize breeders or pathologists. Aflatoxin was discovered in England in 1961 following an epidemic in poultry. By the early 1970s, surveys of agricultural commodities in the USA found that maize produced in the Southeast was especially vulnerable to aflatoxin contamination. Aflatoxin contamination was initially treated as a post-harvest issue, but pre-harvest contamination was proven by 1975. Pre-harvest contamination meant that genetically based host-plant resistance was a possible solution. The potential magnitude of the problem became apparent in 1977 when the southeastern US maize crop suffered epidemic aflatoxin contamination. The first experiment demonstrating the heritability of host-plant resistance to aflatoxin accumulation was published in 1978. These events combined to make breeding for reduced aflatoxin contamination both a high priority and a rational breeding objective. This review surveys the early scientific literature in order to place research on the genetics of aflatoxin accumulation in maize into historical context. It tells the story of how multi-disciplinary research began with veterinary diseases of unknown etiology and resulted in host-plant resistance to a previously minor plant pathogen becoming a central public sector breeding objective.

First report of Fusarium foetens as a mycotoxin producer Abstract Fusarium foetens, a pathogen of Begonia plants, has been recently described as a new fungal species. This Fusarium species causes a destructive vascular wilt disease which leads to the death of the plant. Moreover, Fusarium species are known to produce a huge variety of secondary metabolites such as mycotoxins and phytotoxins. Here, we studied the toxicogenic profile of one F. foetens strain, isolated from maize, employing two methods based on the use of ultra-performance liquid chromatography coupled to mass spectrometry-ion trap-time of flight detection. The mycotoxins beauvericin and fusaric acid were detected in a pure culture of F. foetens. In addition, four fusaric acid analogs (10,11-dihidroxyfusaric acid, hydroxyfusaric acid, dehydrofusaric acid, and a hydroxylated unsaturated fusaric acid analog) were tentatively identified on the basis of their accurate mass and fragmentation patterns. Therefore, these preliminary data indicate that F. foetens isolated from maize is able to produce Fusarium mycotoxins including beauvericin and fusaric acid.

Effects of deoxynivalenol, 3-acetyl-deoxynivalenol and 15-acetyl-deoxynivalenol on parameters associated with oxidative stress in HepG2 cells Abstract This work studied the effect of deoxynivalenol (DON) and two of its acetylated analogs (3-ADON, 15-ADON) on first indicators of oxidative stress status, namely production of reactive oxygen species (ROS) and induction of lipid peroxidation (LPO), in HepG2 cells. HepG2 cells were exposed to different concentrations of the three toxins, either alone or in combinations, for 24, 48, and 72 h. Results of cytotoxicity obtained in HepG2 cells were correlated with the detection of ROS and LPO. This effect was inversely correlated with ROS while directly correlated with LPO for the assayed mycotoxins in individual treatment. Combinations of two toxins containing 15-ADON yielded highest values, while for two-toxin combinations with 3-ADON, the effects were minor. A combination of all three mycotoxins alleviated ROS production and the highest levels in LPO were detected, in association to a final breakdown of adaption of ROS early produced by HepG2. In conclusion, parameters of stress evaluation presented in this study (ROS and LPO), revealed increases in HepG2 cells exposed to DON, 3-ADON, and 15-ADON either individually or combined.

Mycotoxins in poultry feed and feed ingredients in Nigeria Abstract Mycotoxins are toxic secondary fungal metabolites that can negatively affect animal productivity when ingested through feed. In order to assess mycotoxin contamination of poultry feed and feed ingredients vis-a-vis source tracking of feed contamination in Nigeria, 102 samples of feed (n = 30) and feed ingredients (n = 72) were collected from in-house mills of poultry farms across 12 states of Nigeria and analyzed for multiple mycotoxins using LC/MS-MS. One hundred and forty microbial metabolites were detected in the feed and feed ingredients. The most frequent mycotoxin in the feed was fumonisin B1, occurring in 97% of the samples at mean concentration of 1014 μg kg−1. AFB1 occurred in 83% of the feed samples at mean concentration of 74 μg kg−1 and in all feed ingredients except fish meal and other cereals (millet and rice). Feed samples analyzed in this study were contaminated with at least four mycotoxins: aflatoxins and fumonisin co-occurring in 80% of the samples. Peanut cake and maize contributed the most to the levels of aflatoxin and fumonisin, respectively, in the feed. Consequently, there is a need to explore other cereal- and protein-based ingredients for compounding feeds in order to reduce the risk associated with high mycotoxin (e.g. aflatoxin) intake in poultry.