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PubMed1978

Mycotoxins in foodstuffs. XII. The influence of the water activity (a-w) of cakes on the growth of moulds and the formation of mycotoxins.

The influence of various a-w values (high, moderate and low, e.g. 0.92, 0.87, and 0.82) of cakes ("Rührkuchen", made of wheat flour, sugar, eggs, fat and water) on the development and the mycotoxin production of Aspergillus flavus (aflatoxins), A. versicolor (sterigmatocystin), A. ochraceus (ochratoxin A), Penicillium chrysogenum (citrinin) and P. expansum (patulin) was investigated. None of the moulds was able to germinate at an a-w of 0.82. An increase of the water activity resulted in a pronounced growth of the five fungi. Citrinin was not formed on the cakes at all. The yields of the other mycotoxins were not markedly influenced by the a-w (0.92 or 0.87). During the development of the moulds the a-w of the substrate was lowered at first (water requirement during germination and initial growth) and subsequently increased (initiation of the secondary metabolism).

Aspergillus↗
PubMed1998

Studies on toxigenic fungi in roasted foodstuff (salted seed) and halotolerant activity of emodin-producing Aspergillus wentii.

Commercial roasted salted peanuts (3% NaCl), popcorn (1% NaCl), summer-squash (9% NaCl), sunflower (3% NaCl) and wild-melon (3% NaCl) seeds are polluted with fungi, mostly Aspergillus flavus, A. niger, Penicillium chrysogenum, P. corylophilum and Rhizopus stolonifer. Contamination of popcorn with the fungi is about 10 times higher than in the other foods. These fungi, common also on unsalted seeds, are significantly inhibited in seeds (30% moisture content) treated with 9-21% NaCl. The halotolerant A. wentii represents the main fungus recovered from seeds treated by 15-21% NaCl. 9% NaCl stimulated emodin production by A. wentii on peanut and citrinin production by P. chrysogenum on popcorn and sunflower. Aflatoxin, citrinin and emodin production on popcorn persisted up to 15% NaCl. Popcorn is thus strongly susceptible to fungal invasion and toxin pollution. The halotolerance of A. wentii was confirmed by its strong permanent growth in liquid medium at up to 15% NaCl. At 3% NaCl the mycelial growth and nitrogen content increased while the level of emodin and lipid production decreased. CO2 evolution strongly increased at 9-15% NaCl as a characteristic of A. wentii salt tolerance. Emodin inhibited seed viability and the inhibition dose for 50% reduction (LD50) was 65 mg/L for popcorn and 45 mg/L for sunflower.

Aspergillus↗
PubMed1988

Mycotoxin production by molds isolated from 'Greek-style' black olives.

Seventeen mold strains were isolated from 'Greek-style' black olives produced in Morocco. Eight of these isolates were identified as Aspergillus flavus, seven as Aspergillus petrakii, and two as Aspergillus ochraceus Wilhelm. The A. flavus strains were tested for production of aflatoxins B1, B2, G1, and G2; and A. ochraceus and A. petrakii strains were tested for production of ochratoxin, penicillic acid, patulin, and citrinin. The organisms were tested for mycotoxin production on five different substrates, including rice powder-corn steep agar, autoclaved rice, yeast-extract sucrose broth (YES), potato dextrose agar (PDA), and fresh olive paste. All strains of A. flavus produced aflatoxins on all substrates except olive paste and PDA. In PDA, only two strains produced Aflatoxin B1. Five A. ochraceus group isolates produced penicillic acid on one or more of the substrates, but only two out of the five produced penicillic acid on olive paste. None produced ochratoxin, patulin or citrinin. Quantities of aflatoxin B1 produced in rice ranged from 5 to 14 micrograms/g of rice, and of penicillic acid 15-32 micrograms/g of rice. In olive paste, the concentrations of penicillic acid were 11.4 and 30.2 micrograms/g. Biological toxicity of extracts of mold cultures was confirmed using chicken embryos and a microbiological test. Crude extracts of cultures were also tested for mutagenicity using the Salmonella mutagenicity (Ames) Test, and some gave positive mutagenic responses.

Animals↗
PubMedDate not supplied

Quantitative analysis of initiating and promoting activities of five mycotoxins in liver carcinogenesis in rats.

The initiating and promoting activities of several mycotoxins were investigated in F344 male rats. N-2-Fluorenylacetamide (2-FAA) was used as an initiator or a promoter and citrinin, ochratoxin A, sterigmatocystin, (+)rugulosin or patulin was administered at a fixed dose for the same period in the initiation stage or the promotion stage. Partial hepatectomy and administration of carbon tetrachloride (CCl4) were examined to increase the induction of hyperplastic nodules. Ochratoxin A, sterigmatocystin and (+)rugulosin administered in either the initiating or the promoting stage significantly increased the induction of hyperplastic nodules. Citrinin and patulin administered in the initiating stage significantly increased the induction of hyperplastic nodules, but were not effective when administered during the promoting stage. The theoretical classification of the mycotoxins examined is discussed.

Animals↗
PubMed1989

The effect of combined treatment with potassium dichromate and maleic acid on renal function in the rat.

Subthreshold doses of potassium dichromate (K2Cr2O7) have been shown to enhance or potentiate the nephrotoxic effects of mercuric chloride and citrinin. The mechanism of this effect is unknown. The purpose of this study was to examine the phenomenon further by investigating the interaction of K2Cr2O7 with maleic acid, a nephrotoxicant with an action though to be different from those above. Male rats were housed in stainless steel metabolism cages and received a single intraperitoneal injection of maleic acid (300 mg/kg). K2Cr2O7 (10 mg/kg s.c.) or the combination of maleic acid plus K2Cr2O7. While treatment with either agent alone produced only minimal alterations in renal function, the combination of agents produced marked changes in 24 h urine volume, glucose excretion and osmolality. Water consumption was unaffected by the combined treatment. A marked decline in the capacity of renal slices to accumulate both organic anions (p-aminohippurate) and cations (tetraethylammonium) was observed after treatment with both substances. Modifications in renal and hepatic non-protein sulfhydryl content also were observed. These results suggest that K2Cr2O7, as was observed with both mercury and citrinin, enhanced or potentiated the nephrotoxic effects of maleic acid, probably in the proximal tubule, the principal site of glucose reabsorption and organic ion secretion.

Animals↗
PubMed2005

Two primer pairs to detect OTA producers by PCR method.

Fungi contaminating foods and feeds may produce many mycotoxins including ochratoxin A (OTA). Early and rapid detection of potential OTA producing fungi is important to reduce the negative impacts of OTA. In this study, two PCR specific primer pairs, AoLC35-12L/AoLC35-12R and AoOTAL/AoOTAR, were designed from a DNA sequence of a polyketide synthase gene in Aspergillus ochraceus NRRL 3174. On 14 different fungi tested by PCR, AoLC35-12L/AoLC35-12R amplified a unique band from either OTA or citrinin producers while AoOTAL/AoOTAR amplified one PCR product only from A. ochraceus. So these primers could be used to detect both OTA and citrinin producing fungi (AoLC35-12L/AoLC35-12R) or only A. ochraceus (AoOTAL/AoOTAR) from foodstuffs using PCR method.

Amino Acid Sequence↗
PubMed2006

Mycotoxin-induced depletion of intracellular glutathione and altered cytokine production in the human alveolar epithelial cell line A549.

Mould exposure has been associated with asthma and other inflammatory airway conditions. However, cellular effects of inhaled mould components are not well understood. We hypothesised that host defence mechanisms, such as production of cytokines (TGFbeta1, IL-6 and IL-8) and the intracellular antioxidant glutathione (GSH), could be adversely affected by different concentrations of mycotoxins. We studied the effects of citrinin and gliotoxin on lipopolysaccharide (LPS)-stimulated alveolar epithelial cells (A549). Cytokines in cell culture supernatants were analysed by ELISA and levels of GSH were measured by colorimetric (absorbance) determination. We found that GSH decreased in a dose- and time-dependent manner when cells were exposed to citrinin in particular. TGFbeta1 was moderately reduced at low mycotoxin concentrations but elevated at higher sub-toxic concentrations. A tendency for an inverse relationship between TGFbeta1 and GSH levels was observed. IL-6 and IL-8 were not significantly reduced at non-toxic mycotoxin concentrations. Thus, reduced epithelial GSH and TGFbeta1 levels combined with elevated IL-6 and IL-8 levels may result in increased pro-inflammatory activity during mycotoxin exposure. We suggest that this mechanism can contribute to inflammation in mould exposure.

Cell Line↗
PubMed2002

Role of human organic anion transporter 4 in the transport of ochratoxin A.

The purpose of this study was to investigate the characteristics of ochratoxin A (OTA) transport by multispecific human organic anion transporter 4 (hOAT4) using mouse proximal tubule cells stably transfected with hOAT4 (S(2) hOAT4). Immunohistochemical analysis revealed that hOAT4 protein was localized to the apical side of the proximal tubule. S(2) hOAT4 expressed hOAT4 protein in the apical side as well as basolateral side and the cells were cultured on the plastic dish for experiments. S(2) hOAT4 exhibited a time- and concentration-dependent, and a saturable increase in OTA uptake, with an apparent K(m) value of 22.9+/-2.44 microM. The OTA uptakes were inhibited by several substrates for the OATs. Probenecid, piroxicam, octanoate and citrinin inhibited OTA uptake by hOAT4 in a competitive manner (K(i)=44.4-336.4 microM), with the following order of potency: probenecid > piroxicam > octanoate >citrinin. The efflux of OTA by S(2) hOAT4 was higher than that by mock. Addition of OTA resulted in slight decrease in viability of S(2) hOAT4 compared with mock. These results indicate that hOAT4 mediates the high-affinity transport of OTA on the apical side of the proximal tubule, whereas the transport characteristics of OTA are distinct from those by basolateral OATs.

Animals↗
PubMed2006

Mycoflora and multimycotoxin detection in corn silage: experimental study.

Agricultural activities involve the use of crop preservation such as "trench-type" silo, which can sometimes be contaminated by fungi. To investigate the exposure of livestock and farm workers to fungal spores and mycotoxins, a multimycotoxin analysis method has been developed. Six mycotoxins (aflatoxin B1, citrinin, deoxynivalenol, gliotoxin, ochratoxin A, and zearalenone) were quantified by high-performance liquid chromatography coupled to mass spectrometry after solid-phase extraction. An experimental study of fungal species and mycotoxins was conducted in corn silage (Normandy, France) during 9 months of monitoring. The results indicated the recurrence of around 20 different species, with some of them being potentially toxigenic fungi such as Aspergillus fumigatus, Aspergillus parasiticus, Fusarium verticillioides, and Monascus ruber, and the detection of aflatoxin B1 (4-34 ppb), citrinin (4-25 ppb), zearalenone (23-41 ppb), and deoxynivalenol (100-213 ppb). This suggested a possible chronic exposure to low levels of mycotoxins.

Aspergillus↗
PubMedDate not supplied

Occurrence of mycotoxins in raw ingredients used for animal feeding stuffs in the United Kingdom in 1992.

Examination of 330 samples of animal feed ingredients for the presence of a number of mycotoxins has been carried out. These samples were drawn from 186 animal feed mills in the United Kingdom. Aflatoxin B1 was the mycotoxin found most frequently, occurring in most samples of rice bran, maize products, palm kernels and cottonseed, but not in only 3 out of 20 samples of sunflower, in 1 out of 20 samples of soya and in no samples of peas, beans or manioc. Analytical difficulties were met with some combinations of commodity and mycotoxin and all results are uncorrected for recovery. The highest level was detected in a sample of maize gluten: 41 micrograms/kg of aflatoxin B1 (47 micrograms/kg total aflatoxins). Maize products also frequently contained fumonisins B1 and B2 at levels up to nearly 5,000 micrograms/kg in total and zearalenone up to a maximum level of 500 micrograms/kg. Ochratoxin A and citrinin were found in approximately 20% of wheat and barley samples. One sample of barley contained ochratoxin A at a level of 102 micrograms/kg and citrinin at a level of 8 micrograms/kg. Low levels of ochratoxin A also occurred in a few samples of other ingredients: rice bran, palm kernel and beans. Sterigmatocystin at 18 micrograms/kg was found in one sample of organically grown wheat and a trace amount of zearalenone in one sample of manioc. Multi-mycotoxin contamination also occurred, particularly in some samples of maize for which 19 out of 50 samples contained both aflatoxins and fumonisins.

Animal Feed↗
PubMedDate not supplied

High performance liquid chromatography of mycotoxin metabolites in human urine.

Because mycotoxins occur worldwide in grain and grain products, evaluating their effects on the health of the population has become important. The development of a high performance liquid chromatographic (HPLC) procedure was investigated for the analysis of aflatoxin B1, citrinin, and ochratoxin A, from hydrolyzed human urine. Solid-phase extraction (SPE) was used for sample clean-up and concentration. Reversed-phase liquid chromatography (RPLC) was used with fluorescence detection for sample analysis. The presence of aflatoxin B1 was confirmed by converting it to the hemiacetal. With 10 mL of urine, the detection limit for aflatoxin B1 and ochratoxin A was in the high parts per trillion (ppt) and that for citrinin was about 10 ppb.

Aflatoxin B1↗
PubMed2005

Ochratoxin A producing Penicillium verrucosum isolates from cereals reveal large AFLP fingerprinting variability.

AIMS: To examine if molecular amplified fragment length polymorphism (AFLP) fingerprinting of the only ochratoxin A-producing species in European cereals, Penicillium verrucosum, can be used as a method in hazard analysis using critical control points (HACCP). METHODS AND RESULTS: A total of 321 isolates of P. verrucosum were isolated from ochratoxin A-contaminated cereals from Denmark (oats), UK (wheat and barley) and Sweden (wheat). Of these, 236 produced ochratoxin A as determined by thin layer chromatography; 185 ochratoxin A-producing isolates were selected for AFLP fingerprinting. A total of 138 isolates had unique AFLP patterns, whereas 52 isolates could be allocated to small groups containing from two to four isolates with similar AFLP patterns. A total of 155 clones were found among the 185 P. verrucosum isolates, thus 84% of the isolates may represent different genets of P. verrucosum. As the few isolates that were grouped often came from the same farm, and those groups that contained AFLP-identical isolates from different countries were morphotypically different. On single farms up to 35 clones were found. The few groups of ramets from the same genet indicated that a HACCP approach based on clones may require a very large number of AFLP analysis to work in practice, we recommend basing the HACCP approach on the actual species P. verrucosum. A more detailed characterization should rather be based on the profile of species present at different control points, or analysis of the mycotoxins ochratoxin A and citrinin in the isolates. Examination of 86 isolates with HPLC and diode array detection of P. verrucosum showed that 66% produced ochratoxin A, 87% produced citrinin, 92% produced verrucin and 100% produced verrucolone. CONCLUSIONS: Among 184 ochratoxin A-producing Penicillium verrucosum, 155 clonal lineages were indicated by AFLP fingerprinting, indicating a high genetical diversity, yet the species P. verrucosum is phenotypically distinct and valid. SIGNIFICANCE AND IMPACT OF THE STUDY: AFLP fingerprinting of Penicillium verrucosum indicates that genetic recombination takes place in this fungus.

Avena↗
PubMed1979

Toxins of molds from decaying tomato fruit.

Among 27 mold isolates from decaying tomatoes, culture filtrates or ethyl acetate extracts of 8 isolates grown in yeast extract-sucrose medium were markedly toxic (mortality, greater than 50%) to brine shrimp larvae. The toxicity of six of these isolates could be attributed to the presence of citrinin, tenuazonic acid, or T-2 toxin. Ethyl acetate extracts of five Alternaria isolates and one Fusarium isolate were mutagenic for Salmonella typhimurium strains. In ripe tomatoes inoculated with toxin-producing isolates and incubated at 25 degrees C, one Alternaria alternata isolate produced tenuazonic acid in seven of seven tomatoes at levels of up to 106 micrograms/g and alternariol methyl ether in one of the seven tomatoes at 0.8 microgram/g. Another A. alternata isolate produced tenuazonic acid or alternariol methyl ether at much lower levels in only three of seven tomatoes. Patulin and citrinin were produced by a Penicillium expansum isolate at levels of up to 8.4 and 0.76 microgram/g, respectively. In tomatoes incubated at 15 degrees C, a Fusarium sulphureum isolate produced T-2 toxin, HT-2 toxin, and neosolaniol at levels of up to 37.5, 37.8 and 5.6 micrograms/g, respectively. If these mycotoxins are thermostable, they may occur at detectable levels in tomato products whenever partially moldy tomatoes are used as raw material.

Animals↗
PubMed1982

Bacterial bioluminescence as a bioassay for mycotoxins.

The use of bacterial bioluminescence as a toxicological assay for mycotoxins was tested with rubratoxin B, zearalenone, penicillic acid, citrinin, ochratoxin A, PR-toxin, aflatoxin B1, and patulin. The concentrations of mycotoxins causing 50% light reduction (EC50) in Photobacterium phosphoreum were determined immediately and at 5 h after reconstitution of the bacteria from a freeze-dried state. Generally, less toxins were required to obtain an EC50 at 5 h. The effects of the above mycotoxins on bioluminescence were determined after 5, 10, 15, and 20 min of incubation with the bacterial suspensions. The concentration of rubratoxin B necessary to elicit an EC50 increased with time, whereas the concentration of citrinin, penicillic acid, patulin, and PR-toxin necessary decreased with time. There was very little change in the concentration of zearalenone, aflatoxin B1, and ochratoxin A required to elicit an EC50 with time. The bacterial bioluminescence assay was most sensitive to patulin and least sensitive to rubratoxin B.

Biological Assay↗
PubMed2001

Biochemical characterization of ochratoxin A-producing strains of the genus Penicillium.

In order to explore the biochemical scope of ochratoxin A-producing penicillia, we screened 48 Penicillium verrucosum isolates for the production of secondary metabolites. Fungal metabolites were analyzed by high-pressure liquid or gas chromatography coupled to diode array detection or mass spectrometry. The following metabolites were identified: ochratoxins A and B, citrinin, verrucolones, verrucines, anacines, sclerotigenin, lumpidin, fumiquinazolines, alantrypinones, daldinin D, dipodazine, penigequinolines A and B, 2-pentanone, and 2-methyl-isoborneol. By use of average linking clustering based on binary (nonvolatile) metabolite data, the 48 isolates could be grouped into two large and clearly separated groups and a small outlying group of four non-ochratoxin-producing isolates. The largest group, containing 24 isolates, mainly originating from plant sources, included the type culture of P. verrucosum. These isolates produced ochratoxin A, verrucolones, citrinin, and verrucines and had a characteristic dark brown reverse color on yeast extract-sucrose agar medium. Almost all of a group of 20 isolates mainly originating from cheese and meat products had a pale cream reverse color on yeast extract-sucrose agar medium and produced ochratoxin A, verrucolones, anacines, and sclerotigenin. This group included the former type culture of P. nordicum. We also found that P. verrucosum isolates and three P. nordicum isolates incorporated phenylalanine into verrucine and lumpidin metabolites, a finding which could explain why those isolates produced relatively lower levels of ochratoxins than did most isolates of P. nordicum.

Chromatography, Gas↗
PubMedDate not supplied

The role of transport in chemical nephrotoxicity.

Various physiologic factors play a role in determining the extent of chemical-induced nephrotoxicity. One such factor relates to the transport systems that exist in the kidney. Several examples can be given of organic substances that are nephrotoxic only after being transported into renal tubular cells. Some of the cephalosporin antibiotics have been shown to produce proximal tubular necrosis after transport into those cells. Blockade of transport by competitors eliminates or reduces the nephrotoxic response. Citrinin, a secondary product of fungal metabolism, also produces proximal tubular necrosis, but only after transport into proximal tubular cells. Both the cephalosporins and citrinin utilize the organic anion transporter for entry into the cells, a transporter present in adult animals of all species and probably important physiologically for moving metabolic substrates into cells. Various glutathione conjugates (e.g., S-(1,2-dichlorovinyl) glutathione [DCVG]) also are transported into proximal tubular cells with a resulting nephrotoxicity. DCVG utilizes the sodium-dependent transport process that moves glutathione into proximal tubular cells, a process that is inhibited by probenecid. Finally, certain heavy metals also are transported into renal tubular cells. For example, mercuric ion enters proximal cells both from the luminal and peritubular sides and sulfhydryl compounds modify the transport. Movement of mercury from the peritubular side of the cell may be modified by certain organic anions. The characteristics of these mechanisms are less well understood than the mechanisms for the organic compounds.

Animals↗
PubMed2001

[Toxic activity of Penicillium citrinum metabolites on Mus musculus mice].

The aims of this work were to study the influence of secondary metabolites produced by a Penicillium citrinum strain. Mus musculus mice were fed with a diet containing those metabolites to determine both its influence on their development and the pathological and biochemical changes on experimental animals. Male and female including pregnant females, were studied. One group received commercial feed (A.B.) to which the citrinin- producing mould had been added (LF), another received A.B. contaminated with commercial citrinin (LC). The last group received noncontaminated feed (LT). The animals were weighed weekly, and sacrificed after sixty days, so that they could be studied both macro and microscopically. In LF and LT mice, haematological analysis was carried out and hemosiderin was looked for in urine. The diet of the newborne mice, after weaning, was identical to that of their parent. The treated animals (LF and LC) showed morphological alterations, a significant decrease in weight and morphologic alterations and hemosiderin granules were detected in some of their organs. In LT all breeds survived, none of the mice showed neither macro nor microscopic anomalies and had normal biochemical parameters. Fewer breeds in LF survived, male infertility was detected and some of their haematologic parameters were also measurably lower.

English Abstract↗
PubMedDate not supplied

Worldwide occurrence of mycotoxins in foods and feeds--an update.

In a review presented at the first FAO/WHO/UNEP Conference on Mycotoxins in 1977, the occurrence of aflatoxins, zearalenone, ochratoxin A, citrinin, trichothecenes, patulin, penicillic acid, and the ergot alkaloids was indicated to be significant in naturally contaminated foods and feeds. The information presented on aflatoxin contamination greatly exceeded that for all other mycotoxins combined. This study reviews the worldwide levels and occurrence of mycotoxins in various commodities since 1976. Comparatively few countries have lowered the acceptable levels for aflatoxins in susceptible commodities. However, intensified efforts are needed to establish control of aflatoxin levels in the global food supply, particularly in peanuts, tree nuts, corn, and animal feeds. Extensive deoxynivalenol (DON) contamination of grains, especially wheat, was demonstrated. Co-contamination of grains by Fusarium toxins, especially DON and nivalenol, with zearalenone to a lesser extent, was reported. However, more information on co-occurrence of Fusarium toxins in cereals should be developed. When contamination of feeds by ochratoxin A was significant, this toxin occurred in swine kidney and smoked meats in high levels. On the basis of occurrence and/or toxicity, patulin and penicillic acid contamination of foods does not appear to be of real concern. More recent developments suggest, however, that expanded monitoring studies of Alternaria toxins, moniliformin, citrinin, cyclopiazonic acid, penitrem A, and ergot alkaloids are indicated.

Animal Feed↗