Bacillus velezensis as antagonist towards Penicillium roqueforti s.l. in silage: in vitro and in vivo evaluation.

AIMS: The present study was conducted to evaluate the antagonistic effect of Bacillus velezensisNRRL B-23189 towards Penicillium roqueforti s.s. and Penicillium paneum (designated together as P. roqueforti s.l.) in silage conditions. METHODS AND RESULTS: Corn silage conditions were simulated in vitro, and the impact of B. velezensis culture supernatant or cell suspension on P. roqueforti s.l. growth and roquefortine C production was evaluated. The antagonism was promising, but growth of B. velezensis in corn silage infusion was poor. Additionally, an in vivo experiment was carried out with mini-silos containing a mixture of perennial ryegrass and white clover inoculated with P. roqueforti s.l. The applied B. velezensis cell suspension was unsuccessful in reducing P. roqueforti s.l. numbers, but did not compromise the silage acidification. CONCLUSIONS: Although the antagonism observed in vitro was promising, the applied B. velezensis cell suspension could not live up to the expectations in vivo. SIGNIFICANCE AND IMPACT OF THE STUDY: To our knowledge, the present study is the first one evaluating the antagonistic properties of B. velezensis towards toxigenic moulds in silage conditions, offering a good base for further research.

Transcriptome Analysis of Aspergillus flavus Reveals veA-Dependent Regulation of Secondary Metabolite Gene Clusters, Including the Novel Aflavarin Cluster.

The global regulatory veA gene governs development and secondary metabolism in numerous fungal species, including Aspergillus flavus. This is especially relevant since A. flavus infects crops of agricultural importance worldwide, contaminating them with potent mycotoxins. The most well-known are aflatoxins, which are cytotoxic and carcinogenic polyketide compounds. The production of aflatoxins and the expression of genes implicated in the production of these mycotoxins are veA dependent. The genes responsible for the synthesis of aflatoxins are clustered, a signature common for genes involved in fungal secondary metabolism. Studies of the A. flavus genome revealed many gene clusters possibly connected to the synthesis of secondary metabolites. Many of these metabolites are still unknown, or the association between a known metabolite and a particular gene cluster has not yet been established. In the present transcriptome study, we show that veA is necessary for the expression of a large number of genes. Twenty-eight out of the predicted 56 secondary metabolite gene clusters include at least one gene that is differentially expressed depending on presence or absence of veA. One of the clusters under the influence of veA is cluster 39. The absence of veA results in a downregulation of the five genes found within this cluster. Interestingly, our results indicate that the cluster is expressed mainly in sclerotia. Chemical analysis of sclerotial extracts revealed that cluster 39 is responsible for the production of aflavarin.

Dietary mycotoxin exposure and human health risks: A protocol for a systematic review.

BACKGROUND: Mycotoxins are toxic fungal secondary metabolites that contaminate a wide spectrum of essential foods worldwide, such as grain-based products, nuts and spices, causing adverse health effects pertaining to their carcinogenic, nephrotoxic and hepatotoxic nature, among others. AIM: The aim of this systematic review (SR) is to systematically search for, appraise and synthesize primary research evidence to identify what is known about dietary mycotoxin-related health effects and what remains unknown, as well as the uncertainty around findings and the recommendations for the future. SEARCH STRATEGY AND ELIGIBILITY CRITERIA: Search strategies, as well as eligibility criteria were structured according to a predefined PECO (population, exposure, comparison, and outcome) research question and developed in an iterative scoping process. Several bibliographic databases, including Embase, Cochrane Library, Pubmed, Web of Science Core Collection and Scopus, will be searched. Primary research on any measured or modelled dietary exposure to a single or multiple mycotoxins, and adverse human health outcomes (i.e. cancer, non-carcinogenic diseases, and reproductive & developmental adverse outcomes) will be included, and references will be imported into Covidence. In vitro, ex vivo, in silico, animal and review studies, as well as expert's opinions, secondary literature, conference abstracts, presentations, posters, book chapters, dissertations and studies involving non-dietary mycotoxin exposure, will be excluded. STUDY SELECTION: Two independent reviewers will screen titles and abstracts, and review full-texts. Any disagreements will be resolved by a third reviewer based on two-third majority. DATA EXTRACTION: Data from retained eligible studies will be extracted by the principal reviewer, and peer-checked by a second reviewer. STUDY QUALITY ASSESSMENT: Eligible studies will be evaluated for risk of bias (Overall High-Quality Assessment Tool, OHAT) and certainty of evidence (Grading of Recommendations Assessment, Development and Evaluation, GRADE). EVIDENCE SYNTHESIS: A detailed summary of the included studies will be provided within a tabular format and narratively discussed. Heat maps will be constructed to provide information on available knowledge (gaps), and a meta-analysis may be performed based on the variability in predefined PECO elements and depending on the heterogeneity of studies. CONCLUSION: This protocol describes the methodology for the conduct of a SR on mycotoxin-related human health risks, that could guide future research and inform regulatory decisions, as emphasized by the European Commission within the field of regulatory risk assessment for emerging chemicals.

Quantum dot loaded liposomes as fluorescent labels for immunoassay.

Liposomes loaded with water-soluble and water-insoluble quantum dots (QD) were for the first time applied as labels in different heterogeneous immunoassays for the determination of food contaminants, using mycotoxin zearalenone (ZEN) as a model. A great deal of work was devoted to the optimal choice of phospholipids for the liposomes preparation and to the factors which are important for the stability and size of obtained liposomes. Thin-film hydration and reverse-phase evaporation techniques were evaluated in terms of stability of the obtained liposomes and their efficiency for QD loading. Conjugation of liposomes with proteins and the influence of cross-linkers to the nonspecific interaction of the obtained liposomes with the surface of microtiter plates and cartridges were investigated and 3-(2-pyridyldithio)propionic acid N-hydroxysuccinimide ester was found as the optimal cross-linker. The limits of detection (LOD) for ZEN of fluorescence-labeled immunosorbent assays were 0.6 µg kg(-1), 0.08 µg kg(-1), and 0.02 µg kg(-1), using QD, liposomes loaded with water-soluble QD, and water-insoluble QD, respectively. Similarly, the developed qualitative on-site tests using the different QD labels and taking into account the EU maximum residues level for ZEN in unprocessed cereals showed cutoff levels of 100, 50, and 20 µg kg(-1).

Liposomes loaded with quantum dots for ultrasensitive on-site determination of aflatoxin M1 in milk products.

A quantitative fluorescence-labeled immunosorbent assay and qualitative on-site column tests were developed for the determination of aflatoxin M1 in milk products. The use of liposomes loaded with quantum dots as a label significantly increased the assay sensitivity by encapsulating multiple quantum dots in a single liposome and, therefore, amplifying the analytical signal. Two different techniques were compared to obtain aflatoxin-protein conjugates, used for further coupling with the liposomes. The influence of nonspecific interactions of the liposome-labeled conjugates obtained with the surface of microtiter plates and column cartridges was evaluated and discussed. The limit of detection for fluorescence-labeled immunosorbent assay was 0.014 µg kg(-1). For qualitative on-site tests, the cutoff was set at 0.05µg kg(-1), taking into account the EU maximum level for aflatoxin M1 in raw milk, heat-treated milk, and milk for the manufacture of milk-based products. The direct addition of labeled conjugate to the milk samples resulted in an additional decrease of analysis time. An intralaboratory validation was performed with sterilized milk and cream samples artificially spiked with aflatoxin M1 at concentrations less than, equal to and greater than the cutoff level. It is shown that milk products can be analyzed without any sample preparation, just diluted with the buffer. The rates for false-positive and false-negative results were below 5% (2.6% and 3.3%, respectively).

Quantum dot based rapid tests for zearalenone detection.

Three different kinds of immunosorbent assays with luminescence detection were developed for the determination of zearalenone (ZEN), a secondary toxic metabolite of Fusarium fungi. CdSe/ZnS core/shell quantum dots (QDs) were used as a label in quantitative micro-well plate immunoassays (fluorescent-labeled immunosorbent assay, FLISA) and in qualitative column test methods. As carriers for QD-based column tests, sepharose gel (for covalent binding of antibody) and polyethylene frits (for physical absorption of antibody) were used and compared. The application of QDs as a label resulted in a fourfold decrease in the IC(50) value with FLISA (0.1 ng mL(-1)) with a detection limit of 0.03 ng mL(-1) when compared with the traditional immunosorbent assay which makes use of horseradish peroxidase as the enzyme label. The cutoff levels for both qualitative column test methods were selected based on the maximum level for ZEN in unprocessed cereals established by the European Commission (100 µg kg(-1)) as 5 ng mL(-1) taking into account extraction and dilution. The different developed immumoassays were tested for ZEN determination in raw wheat samples. As a confirmatory method, liquid chromatography coupled to tandem mass spectrometry was used. The obtained results allow using FLISA and both qualitative column test methods for the analysis of analytes with very low established maximum limits, even in very complicated food matrices, owing to the high dilution of the sample extract.

Multiplex flow-through immunoassay formats for screening of mycotoxins in a variety of food matrices.

Two multi-analyte flow-through immunoassay formats for rapid detection of mycotoxins in a variety of food matrices (peanut cake, maize, and cassava flour) were developed and evaluated. The selected food matrices are typical staple foods and export products for most low-income communities around the world. The assay formats included gel-based and membrane-based flow-through assays and were based on the principle of indirect enzyme-linked immunosorbent assay. Using the same immunoreagents, the performance characteristics of both assays were compared. To the best of our knowledge, this is the first report on such a comparison. The gel-based format was developed to screen for ochratoxin A, fumonisin B(1), deoxynivalenol, and zearalenone detection at cut-off values of 3, 1,250, 1,000, and 200 µg kg(-1), respectively, while the membrane-based format can be used to screen ochratoxin A, aflatoxin B(1,) deoxynivalenol, and zearalenone at the following cut-offs: 3, 5, 700, and 175 µg kg(-1), respectively. The applicability of these assay formats was demonstrated by evaluating the performance characteristics of both tests through performing multiple experiments on different days. Both assays were further evaluated by analyzing naturally contaminated samples in the laboratory and also in the field under tropical conditions (Cameroon, West Africa). The false-negative rate with both formats was less than 5%, which is in good agreement with Commission Decision 2002/657/EC regarding the performance of analytical methods intended for screening purposes.

Efficacy and safety testing of mycotoxin-detoxifying agents in broilers following the European Food Safety Authority guidelines.

Contamination of feeds with mycotoxins is a worldwide problem and mycotoxin-detoxifying agents are used to decrease their negative effect. The European Food Safety Authority recently stated guidelines and end-points for the efficacy testing of detoxifiers. Our study revealed that plasma concentrations of deoxynivalenol and deepoxy-deoxynivalenol were too low to assess efficacy of 2 commercially available mycotoxin-detoxifying agents against deoxynivalenol after 3 wk of continuous feeding of this mycotoxin at concentrations of 2.44±0.70 mg/kg of feed and 7.54±2.20 mg/kg of feed in broilers. This correlates with the poor absorption of deoxynivalenol in poultry. A safety study with 2 commercially available detoxifying agents and veterinary drugs showed innovative results with regard to the pharmacokinetics of 2 antibiotics after oral dosing in the drinking water. The plasma and kidney tissue concentrations of oxytetracycline were significantly higher in broilers receiving a biotransforming agent in the feed compared with control birds. For amoxicillin, the plasma concentrations were significantly higher for broilers receiving an adsorbing agent in comparison to birds receiving the biotransforming agent, but not to the control group. Mycotoxin-detoxifying agents can thus interact with the oral bioavailability of antibiotics depending on the antibiotic and detoxifying agent, with possible adverse effects on the health of animals and humans.

Determination of fumonisins B1 and B2 in corn by LC/MS with immunoaffinity column cleanup: interlaboratory study.

An interlaboratory validation study was conducted to establish the method performance characteristics of an immunoaffinity column (IAC) cleanup procedure followed by LC/MS for the determination of fumonisins B1 (FB1) and B2 (FB2) and combined FB1 + FB2 in corn. The test portion is extracted with acetonitrile-methanol-water (25 + 25 + 50). The extract is filtered, diluted with phosphate-buffered saline solution, and applied to an IAC. FB1 and FB2 are removed with methanol, followed by water, then directly determined by RPLC with MS detection using selected-ion monitoring of two characteristic ions in each case. Naturally contaminated corn samples were milled to a fine powder and mixed to produce three samples with target levels of combined FB1 + FB2 ranging from 350 to 4000 microg/kg. Of 15 initially participating laboratories, two failed to report results and another did not follow the prescribed method. Thus, valid results were obtained from 12 participants located in 11 countries. Statistical analysis of the results produced RSDr values of 4.6-11.9, 1.9-12.6, and 1.4-11.5% for FB1, FB2, and combined FB1 + FB2, respectively; the corresponding RSDR values were 19.8-23.8, 18.2-25.5, and 18.8-23.2%. The three concentration levels of combined FB1 + FB2 were 534, 1194, and 1954 microg/kg. HorRat values for r and R were all < 2.0, indicating that the method is suitable as a regulatory method for the enforcement of European Union limits for fumonisins in corn.

Burden of disease associated with dietary exposure to carcinogenic aflatoxins in Portugal using human biomonitoring approach.

Human biomonitoring is an important tool to assess human exposure to chemicals, contributing to describe trends of exposure over time and to identify population groups that could be under risk. Aflatoxins are genotoxic and carcinogenic food contaminants causing hepatocellular carcinoma, the third leading cause of cancer deaths worldwide. In Portugal, scarce data are available regarding exposure to aflatoxins and no previous study used human biomonitoring data to comprehensively characterize the associated burden of disease. 24 h urine and first-morning urine paired samples were collected by 94 participants and were analyzed by liquid chromatography-tandem mass spectrometry for the quantitative determination of aflatoxins (B1, B2, G1, G2 and M1). Deterministic and probabilistic models were developed to assess the Portuguese exposure to aflatoxins and to estimate the health impact of this exposure, estimating the attributed Disability-Adjusted Life Years (DALYs). Aflatoxins were detected in a maximum of 13% (AFB1), 16% (AFB2), 1% (AFG1), 2% (AFG2) and 19% (AFM1) of the urine samples. Data obtained through the probabilistic approach revealed an estimated mean probable daily intake of 13.43 ng/kg body weight per day resulting in 0.13 extra cases of hepatocellular carcinoma, corresponding to mean annual DALYs of 172.8 for the Portuguese population (10291027 inhabitants). The present study generated for the first time and within a human biomonitoring study, reliable and crucial data to characterize the burden associated to the exposure to aflatoxins of the Portuguese population. The obtained results constitute an imperative support to risk managers in the establishment of preventive policy measures that contribute to ensure public health protection.

Natural occurrence of mycotoxins in maize and sorghum in Togo.

Mycotoxins are fungal secondary metabolites frequently affecting agronomical crops and consequently imposing a major challenge for food safety and public health. In this study, a total of 67 raw cereals (55 maize and 12 sorghum) were collected from the market of Togo. The samples were investigated on the occurrence of 21 mycotoxins using state-of-the-art high-performance liquid chromatography coupled to tandem mass spectrometry (HPLC-MS/MS). The most frequent occurring mycotoxins were fumonisins (88 and 67% for maize and sorghum respectively) with concentrations ranging from 101 to 1838 µg/kg for maize and 81.5 to 361 µg/kg for sorghum, respectively. Aflatoxin B1 was detected in 38% of the maize samples with maximum contamination levels of 256 µg/kg, and 25% of the sorghum samples (range 6-16 µg/kg). The concentrations of aflatoxins were high in maize, with some cases exceeding the maximum legislative limits (EU) for unprocessed maize placed on the market. In addition to these high contamination levels, the co-occurrence of three classes of mycotoxins (i.e., aflatoxins, fumonisins, and trichothecenes) was observed in this study. For the first time, the multi-mycotoxins occurrence in agronomical crops in Togo was reported.

Exposure assessment of Portuguese population to multiple mycotoxins: The human biomonitoring approach.

Mycotoxins constitute a relevant group of food contaminants with several associated health outcomes such as estrogenic, immunotoxic, nephrotoxic and teratogenic effects. Although scarce data are available in Portugal, human biomonitoring studies have been globally developed to assess the exposure to mycotoxins at individual level. In order to overcome this lack of data, the present study concerned the analysis of mycotoxins in 24h urine and first-morning urine paired samples from 94 participants enrolled within the scope of the National Food, Nutrition, and Physical Activity Survey of the Portuguese General Population (2015-2016). Following a salt-assisted matrix extraction, urine samples were analysed by liquid chromatography-mass spectrometry for the simultaneous determination of 37 urinary mycotoxins' biomarkers and data obtained used to estimate the probable daily intake as well as the risk characterization applying the Hazard Quotient approach. Results revealed the exposure of Portuguese population to zearalenone, deoxynivalenol, ochratoxin A, alternariol, citrinin and fumonisin B1 through the quantification in 24h urine and first-morning urine paired samples. Risk characterization data revealed a potential concern to some reported mycotoxins since the reference intake values were exceeded by some of the considered participants. Alternariol was identified for the first time in urine samples from a European country; however, risk characterization was not performed due to lack of reference intake value. These results confirmed mycotoxins as part of the human exposome of the Portuguese population reinforcing the need for further studies regarding the determinants of exposure.

Novel gel-based rapid test for non-instrumental detection of ochratoxin A in beer.

A rapid easy-to-use immunoassay was optimised for the non-instrumental detection of ochratoxin A (OTA) in beer. The analytical method involves preconcentration on the immunoaffinity layer inside a column followed by direct competitive ELISA detection in the same layer. The visual cut-off value, i.e. the lowest OTA concentration resulting in no colour development, was 0.2 microg L(-1). Assay validation was performed using samples spiked with OTA. Thirty-seven naturally contaminated samples were screened with the gel-based method developed and no false-negative results were obtained. The method described offers a simple, rapid and cost-effective screening tool, thus contributing to better health protection of consumers.

Processing of mycotoxin contaminated waste streams through anaerobic digestion.

Food and feed stocks heavily contaminated with mycotoxins are rendered unfit for consumption and therefore discarded as waste. Due to the lack of guidelines and in accordance with the prudent avoidance principle, these waste streams are often incinerated. For better valorization, these streams could be used as input for anaerobic digestion. However, the degradation of multiple mycotoxins during anaerobic digestion and their effect on the methane production is currently unknown. In batch tests spiked with mycotoxins, aflatoxin B1, ochratoxin A, deoxynivalenol, zearalenone and T-2 toxin were degraded for more than 90%. For mesophile and thermophile digestion respectively, fumonisin B1 was degraded for 70% and 85%, and most ergot alkaloids for 64% and 98%. Neither biogas production, nor methane production were influenced by the presence of the mycotoxins. Subsequently, semi-continuous reactors fed with contaminated maize resulted in more than 99% degradation for all mycotoxins after 1.8 hydraulic retention time with stable biogas production and process parameters. This study shows that mycotoxin contaminated organic waste can be safely valorized to methane while the digestate is void of mycotoxin residues.

Analysis of alternariol and alternariol monomethyl ether in foodstuffs by molecularly imprinted solid-phase extraction and ultra-high-performance liquid chromatography tandem mass spectrometry.

Molecularly imprinted porous polymer microspheres selective to Alternaria mycotoxins, alternariol (AOH) and alternariol monomethyl ether (AME), were synthesized and applied to the extraction of both mycotoxins in food samples. The polymer was prepared using 4-vinylpiridine (VIPY) and methacrylamide (MAM) as functional monomers, ethylene glycol dimethacrylate (EDMA) as cross-linker and 3,8,9-trihydroxy-6H-dibenzo[b,d]pyran-6-one (S2) as AOH surrogate template. A molecularly imprinted solid phase extraction (MISPE) method has been optimized for the selective isolation of the mycotoxins from aqueous samples coupled to HPLC with fluorescence (λex=258nm; λem=440nm) or MS/MS analysis. The MISPE method was validated by UPLC-MS/MS for the determination of AOH and AME in tomato juice and sesame oil based on the European Commission Decision 2002/657/EC. Method performance was satisfactory with recoveries from 92.5% to 106.2% and limits of quantification within the 1.1-2.8µgkg-1 range in both samples.

Evaluation Of Mycotoxin Content In Soybean (Glycine max L.) Grown In Rwanda.

Soybean is a critical food and nutritional security crop in Rwanda. Promoted by the Rwandan National Agricultural Research System for both adults and as an infant weaning food, soybean is grown by approximately 40% of households. Soybean may be susceptible to the growth of mycotoxin-producing moulds; however, data has been contradictory. Mycotoxin contamination is a food and feed safety issue for grains and other field crops. This study aimed to determine the extent of mycotoxin contamination in soybean, and to assess people's awareness on mycotoxins. A farm-level survey was conducted in 2015 within three agro-ecological zones of Rwanda suitable for soybean production. Soybean samples were collected from farmers (n=300) who also completed questionnaires about pre-and post-harvest farm practices, and aflatoxin awareness. The concentration of total aflatoxin in individual soybean samples was tested by enzymelinked immunosorbent assay (ELISA) using a commercially-available kit. Other mycotoxins were analyzed using liquid chromatography-mass spectrometry (LCMS/MS) on 10 selected sub samples. Only 7.3% of the respondents were aware of aflatoxin contamination in foods, but farmers observed good postharvest practices including harvesting the crop when the pods were dry. Using enzyme-linked immunosorbent assay (ELISA), only one sample had a concentration (11 µg/kg) above the most stringent EU maximum permitted limit of 4 µg/kg. Multi-mycotoxins liquid chromatography-mass spectrometry (LC-MS/MS) results confirmed that soybeans had low or undetectable contamination; only one sample contained 13µg/kg of sterigmatocystine. The soybean samples from Rwanda obtained acceptably low mycotoxin levels. Taken together with other studies that showed that soybean is less contaminated by mycotoxins, these results demonstrate that soybean can be promoted as a nutritious and safe food. However, there is a general need for educating farmers on mycotoxin contamination in food and feed to ensure better standards are adhered to safeguard the health of the consumers regarding these fungal secondary metabolites.

Influence of mycotoxin binders on the oral bioavailability of tylosin, doxycycline, diclazuril, and salinomycin in fed broiler chickens.

The presence of mycotoxins in broiler feed can have deleterious effects on the wellbeing of the animals and their performance. Mycotoxin binders are feed additives that aim to adsorb mycotoxins in the intestinal tract and thereby prevent the oral absorption of the mycotoxin. The simultaneous administration of coccidiostats and/or antimicrobials with mycotoxin binders might lead to a reduced oral bioavailability of these veterinary medicinal products. This paper describes the influence of 3 mycotoxin binders (i.e., clay 1 containing montmorillonite, mica, and feldspars; clay 2 containing montmorillonite and quartz; and yeast 1 being a modified glucomannan fraction of inactivated yeast cells) and activated carbon on the oral bioavailability and pharmacokinetic parameters of the antimicrobials doxycycline and tylosin, and the coccidiostats diclazuril and salinomycin. A feeding study with 40 15 day-old broilers was performed evaluating the effects of long-term feeding 2 g mycotoxin binder/kg of feed. The birds were randomly divided into 5 groups of 8 birds each, i.e., a control group receiving no binder and 4 test groups receiving either clay 1, clay 2, yeast 1, or activated carbon mixed in the feed. After 15 d of feeding, both the control and each test group were administered doxycycline, tylosin, diclazuril, and salinomycin, consecutively, respecting a wash-out period of 2 to 3 d between each administration. The 4 medicinal products were dosed using a single bolus administration directly in the crop. After each bolus administration, blood was collected for plasma analysis and calculation of the main pharmacokinetic parameters and relative oral bioavailability (F = area under the plasma concentration-time curve (AUC0-8 h) in the test groups/AUC0-8 h in the control group)*100). No effects were observed of any of the mycotoxin binders on the relative oral bioavailability of the coccidiostats (i.e., F between 82 and 101% and 79 and 93% for diclazuril and salinomycin, respectively). Also, no significant effects could be noticed of any of the mycotoxin binders on the relative oral bioavailability of the antimicrobials doxycycline and tylosin (i.e., F between 67 and 83% and between 43 and 104%, respectively).

Immunocompetence in organically fed finishing pigs: effect of corn cob mix.

Two consecutive experiments were performed to evaluate the effects on the immune response of corn cob mix (CCM) in an organic pig diet. The immunoglobulin (Ig) M, IgA and IgG responses against an intramuscularly injected model antigen, bovine thyroglobulin, were used as indicator. The experiments were performed in an organic barn with nine pens of four crossbred pigs (two barrows and two sows) from 45 kg to slaughter. In the first experiment, the organic concentrate was mixed with organic CCM-silage to obtain three concentrate: CCM ratios of 100:0, 80:20 and 60:40 (w:w). In the second experiment, three concentrates were produced to obtain diets with equal nutrient levels on a dry matter basis after 0%, 20% and 40% CCM inclusion. Higher inclusion rates of CCM in the ration were accompanied by lower thyroglobulin-specific IgG responses. These effects could not be attributed to one specific component of the CCM, such as fatty acid composition, although there was a degree of correlation with lower vitamin A concentrations. Mycotoxin concentrations were absent or minimal. The study indicated that dietary ingredient composition may affect immunocompetence.

Aphids transform and detoxify the mycotoxin deoxynivalenol via a type II biotransformation mechanism yet unknown in animals.

Biotransformation of mycotoxins in animals comprises phase I and phase II metabolisation reactions. For the trichothecene deoxynivalenol (DON), several phase II biotransformation reactions have been described resulting in DON-glutathiones, DON-glucuronides and DON-sulfates made by glutathione-S-transferases, uridine-diphosphoglucuronyl transferases and sulfotransferases, respectively. These metabolites can be easily excreted and are less toxic than their free compounds. Here, we demonstrate for the first time in the animal kingdom the conversion of DON to DON-3-glucoside (DON-3G) via a model system with plant pathogenic aphids. This phase II biotransformation mechanism has only been reported in plants. As the DON-3G metabolite was less toxic for aphids than DON, this conversion is considered a detoxification reaction. Remarkably, English grain aphids (Sitobion avenae) which co-occur with the DON producer Fusarium graminearum on wheat during the development of fusarium symptoms, tolerate DON much better and convert DON to DON-3G more efficiently than pea aphids (Acyrthosiphon pisum), the latter being known to feed on legumes which are no host for F. graminearum. Using a non-targeted high resolution mass spectrometric approach, we detected DON-diglucosides in aphids probably as a result of sequential glucosylation reactions. Data are discussed in the light of an eventual co-evolutionary adaptation of S. avenae to DON.

Monitoring of polychlorinated biphenyls in Belgian human adipose tissue samples.

Polychlorinated biphenyls (PCBs) were monitored in Belgian human adipose tissue samples from deceased individuals (n=100). Their mean age was 52, ranging from 2 to 91 years. There were 57 men and 43 women. Other known variables were date of autopsy and place of residence. No information about diet or occupation was available. The seven marker congeners PCB 28, 52, 101, 118, 138, 153 and 180 were analysed in the samples with a GC-MS/MS method validated according to Commission Decision 2002/657/EC. Extracted fat was cleaned-up over a glass column filled with n-hexane, acid silica, deactivated alumina and anhydrous sodium sulfate. The whole procedure was subjected to a rigorous quality control programme with retention times, ion chromatograms and intensity ratios of the monitored product ions as identification criteria. The total PCB concentration ranged between 10 and 1640 ng g-1 fat, with a mean value of 658 ng g-1 fat. In the age groups of 0-9 (n=1), 10-19 (n=4), 20-29 (n=11), 30-39 (n=13), 40-49 (n=15), 50-59 (n=14), 60-69 (n=14), 70-79 (n=20), 80-89 (n=6) and 90-99 (n=2), the mean total PCB concentrations were 10, 134, 253, 445, 557, 687, 807, 962, 959, and 1191 ng g-1 fat, respectively. So, there was an increase of PCB body burden with age. For the male subjects (n=57; mean age of 53) the mean total PCB concentration was 633 ng g-1 fat. For the female subjects (n=43; mean age of 52) it was 690 ng g-1 fat. There was no significant sex-related difference in the concentrations of marker PCBs.