Occurrence, prevention and remediation of toxigenic fungi and mycotoxins in silage: a review.

Ruminants are considered to be less sensitive towards mycotoxins than monogastric animals because rumen microbiota have mycotoxin-detoxifying capacities. Therefore the effect of mycotoxins towards ruminants has been studied to a lesser extent compared with monogastric animals. Worldwide, a high proportion of the ruminant diet consists of silages made of forage crops (i.e. all parts of the crop above the stubble are harvested). In practice, silages are often contaminated with multiple mycotoxins. Exposure to a cocktail of mycotoxins can hamper animal production and have severe health consequences. In this article the different aspects associated with mycotoxin contamination of silage are reviewed 'from seed to feed'. An overview is given on the occurrence of toxigenic fungal species and their concomitant mycotoxins in forage crops before and after ensiling. The mycotoxin load of visually non-mouldy samples and mouldy hot spots within the same silo is also compared. Subsequently, this review delves into different problem-solving strategies. A logical first step is prevention of mould growth and mycotoxin production in the field, during harvest and during ensiling. If prevention should fail, several remediation strategies are available. These are listed, mainly focusing on the possibilities of microbial degradation of mycotoxins in vivo in silage. © 2015 Society of Chemical Industry.

Prevalence of BRCA1/2 mutations in sporadic breast/ovarian cancer patients and identification of a novel de novo BRCA1 mutation in a patient diagnosed with late onset breast and ovarian cancer: implications for genetic testing.

In order to adequately evaluate the clinical relevance of genetic testing in sporadic breast and ovarian cancer patients, we offered comprehensive BRCA1/2 mutation analysis in patients without a family history for the disease. We evaluated the complete coding and splice site regions of BRCA1/2 in 193 sporadic patients. In addition, a de novo mutation was further investigated with ultra deep sequencing and microsatellite marker analysis. In 17 patients (8.8%), a deleterious germline BRCA1/2 mutation was identified. The highest mutation detection ratio (3/7 = 42.9%) was obtained in sporadic patients diagnosed with breast and ovarian cancer after the age of 40. In 21 bilateral breast cancer patients, two mutations were identified (9.5%). Furthermore, 140 sporadic patients with unilateral breast cancer were investigated. Mutations were only identified in patients diagnosed with breast cancer before the age of 40 (12/128 = 9.4% vs. 0/12 with Dx > 40). No mutations were detected in 17 sporadic male breast cancer and 6 ovarian cancer patients. BRCA1 c.3494_3495delTT was identified in a patient diagnosed with breast and ovarian cancer at the age of 52 and 53, respectively, and was proven to have occurred de novo at the paternal allele. Our study shows that the mutation detection probability in specific patient subsets can be significant, therefore mutation analysis should be considered in sporadic patients. As a consequence, a family history for the disease and an early age of onset should not be used as the only criteria for mutation analysis of BRCA1/2. The relatively high mutation detection ratio suggests that the prevalence of BRCA1/2 may be underestimated, especially in sporadic patients who developed breast and ovarian cancer. In addition, although rare, the possibility of a de novo occurrence in a sporadic patient should be considered.

Development of an in vitro gastro-intestinal pig model to screen potential detoxifying agents for the mycotoxin deoxynivalenol.

Deoxynivalenol (DON) is a type B trichothecene mycotoxin with worldwide high incidence in feed which is produced by Fusarium species. Strategies are needed to eliminate its health risk for livestock and to minimise its economic impact on production. In order to assess the efficacy of potential physical, chemical and biological DON detoxifying agents, a good in vitro model is necessary to perform a fast and high-throughput screening of new compounds before in vivo trials are set up. In this paper, an in vitro model was developed to screen potential commercial products for DON degradation and detoxification. Contaminated feed with potential detoxifying agents are first applied to a simulated gastrointestinal tract (GIT) of a pig, after which detoxification is assessed through a robust, inexpensive and readily applicable Lemna minor L. aquatic plant bioassay which enables evaluation of the residual toxicity of possible metabolites formed by DON detoxifying agents. The GIT simulation enables taking matrix and incubation parameters into account as they can affect the binding, removal or degradation of DON. One product could reduce DON in feed in the GIT model for almost 100% after 6 h. DON metabolites were tentatively identified with LC-MS/MS. This GIT simulation coupled to a detoxification bioassay is a valuable model for in vitro screening and assessing compounds for DON detoxification, and could be expanded towards other mycotoxins.

Bacterial Enrichment Cultures Biotransform the Mycotoxin Deoxynivalenol into a Novel Metabolite Toxic to Plant and Porcine Cells.

The mycotoxin deoxynivalenol (DON), produced in wheat, barley and maize by Fusarium graminearum and Fusarium culmorum, is threatening the health of humans and animals. With its worldwide high incidence in food and feed, mitigation strategies are needed to detoxify DON, maintaining the nutritional value and palatability of decontaminated commodities. A promising technique is biological degradation, where microorganisms are used to biotransform mycotoxins into less toxic metabolites. In this study, bacterial enrichment cultures were screened for their DON detoxification potential, where DON and its potential derivatives were monitored. The residual phytotoxicity was determined through a bioassay using the aquatic plant Lemna minor L. Two bacterial enrichment cultures were found to biotransform DON into a still highly toxic metabolite for plants. Furthermore, a cytotoxic effect was observed on the cellular viability of intestinal porcine epithelial cells. Through liquid chromatography high-resolution mass spectrometry analysis, an unknown compound was detected, and tentatively characterized with a molecular weight of 30.0 Da (i.e., CH2O) higher than DON. Metabarcoding of the subsequently enriched bacterial communities revealed a shift towards the genera Sphingopyxis, Pseudoxanthomonas, Ochrobactrum and Pseudarthrobacter. This work describes the discovery of a novel bacterial DON-derived metabolite, toxic to plant and porcine cells.

Microbial Detoxification of Deoxynivalenol (DON), Assessed via a Lemna minor L. Bioassay, through Biotransformation to 3-epi-DON and 3-epi-DOM-1.

Mycotoxins are toxic metabolites produced by fungi. To mitigate mycotoxins in food or feed, biotransformation is an emerging technology in which microorganisms degrade toxins into non-toxic metabolites. To monitor deoxynivalenol (DON) biotransformation, analytical tools such as ELISA and liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) are typically used. However, these techniques do not give a decisive answer about the remaining toxicity of possible biotransformation products. Hence, a bioassay using Lemna minor L. was developed. A dose-response analysis revealed significant inhibition in the growth of L. minor exposed to DON concentrations of 0.25 mg/L and higher. Concentrations above 1 mg/L were lethal for the plant. This bioassay is far more sensitive than previously described systems. The bioassay was implemented to screen microbial enrichment cultures, originating from rumen fluid, soil, digestate and activated sludge, on their biotransformation and detoxification capability of DON. The enrichment cultures originating from soil and activated sludge were capable of detoxifying and degrading 5 and 50 mg/L DON. In addition, the metabolites 3-epi-DON and the epimer of de-epoxy-DON (3-epi-DOM-1) were found as biotransformation products of both consortia. Our work provides a new valuable tool to screen microbial cultures for their detoxification capacity.

Biodegradation of Mycotoxins: Tales from Known and Unexplored Worlds.

Exposure to mycotoxins, secondary metabolites produced by fungi, may infer serious risks for animal and human health and lead to economic losses. Several approaches to reduce these mycotoxins have been investigated such as chemical removal, physical binding, or microbial degradation. This review focuses on the microbial degradation or transformation of mycotoxins, with specific attention to the actual detoxification mechanisms of the mother compound. Furthermore, based on the similarities in chemical structure between groups of mycotoxins and environmentally recalcitrant compounds, known biodegradation pathways and degrading organisms which hold promise for the degradation of mycotoxins are presented.