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PubMed2000

Induction of unscheduled DNA synthesis in primary human urothelial cells by the mycotoxin ochratoxin A.

Ochratoxin A (OTA) is a widespread contaminant in human staple food. Exposure of humans to this mycotoxin is a matter of concern because OTA is a known rodent carcinogen. As the urothelium is one target tissue of this mycotoxin, primary cultured human urothelial cells (HUC) from adults and children were used to analyze the induction of unscheduled DNA synthesis (UDS) by OTA. HUC were isolated from the ureters or renal pelves of two nephrectomized adults and of two children with ureteropelvic junction stenosis and cultured under serum-free conditions. After a confluency of 70-80% was reached, cell proliferation was suppressed by arginine-deficient medium (ADM), and UDS was assessed autoradiographically by 3H-thymidine incorporation upon exposure to OTA (10-2000 nM), ethyl methanesulfonate (EMS, 5 mM, positive control), or dimethyl sulfoxide (DMSO, 0.2%, solvent control). In control cultures the level of UDS was low. Exposure to EMS resulted in an induction of UDS (2-to 5-fold compared to control), thus allowing the sensitive detection of repair resulting from induction of DNA lesions in all four specimens, and demonstrating that repair of EMS-induced DNA lesions can take place under the chosen culture conditions. In two HUC cultures derived from adults, a significant induction of UDS was observed in the concentration range of 50-500 nM OTA. The highest fraction of cells in repair (CIR) was found at 50 nM OTA for the HUC from the older male (50% CIR). The maximum response in the other specimens from the adult female and the 7-year-old boy were seen at OTA concentrations of 500 and 250 nM, respectively. In contrast to all other specimens, no significant induction of UDS by OTA was found in the HUC cultures derived from an infant's urothelium. Signs of cytotoxicity were observed above 500 nM OTA in all cultures. The varying susceptibility toward OTA observed in vitro may hint at varying predispositions of individuals in vivo.

Adult↗

PubMed1987

Flavonoids protect against T-2 mycotoxins both in vitro and in vivo.

Quercetin was able to reduce the cytotoxic effect of T-2 mycotoxin on cultured murine thymocytes. When given to mice immediately before challenge with T-2 mycotoxins, quercetin significantly reduced mortality.

Animals↗

PubMed1995

Antiviral activity of trichothecene mycotoxins (deoxynivalenol, fusarenon-X, and nivalenol) against herpes simplex virus types 1 and 2.

The effect of trichothecene mycotoxins, deoxynivalenol (DON), fusarenon-X (FX) and nivalenol (NIV), on plaque formation of herpes simplex virus types 1 and 2 (HSV-1 and HSV-2) in HEp-2 cells was examined. The 50% effective concentrations (EC50) of DON, FX, and NIV for HSV-1 plaque formation were 160, 56, and 120 ng/ml, respectively. Those for HSV-2 plaque formation were 94, 26, and 50 ng/ml, respectively. These three mycotoxins showed about 2-fold higher selectivity to HSV-2 than to HSV-1. Plaque formation of HSV-1 was not inhibited with trichothecenes at concentrations completely inhibiting plaque formation when cells were treated during virus adsorption period or 15 hr before infection. These results indicate that trichothecenes affect replication of HSV-1 after virus adsorption, but not before or during virus adsorption to the host cells.

Cell Line↗

PubMed2005

Metal ions modulate gene expression and accumulation of the mycotoxins aflatoxin and zearalenone.

AIMS: To determine the modulating action of some metal ions (Zn+2, Fe+2, Cu+2) on gene expression of enzymes related to fungal growth and accumulation of the mycotoxins aflatoxin and zearalenone. METHODS AND RESULTS: The effect of the metal ions, as single or mixed treatments, was observed in submerged cultures of toxigenic Aspergillus flavus or Fusarium graminearum, which produce the mycotoxins aflatoxin or zearalenone, respectively. The enzyme-linked immunosorbent assay results showed that the single metals Zn+2 or Cu+2 stimulated aflatoxin accumulation while Cu+2 or Fe+2 stimulated zearalenone in fungal cultures. Single Zn+2 treatment also affected conidial differentiation and pigmentation. A cDNA suppression subtractive library was also produced and followed by sequencing of potential metal treatment-specific clones, thus determining induced genes. The genes uncovered included enzymes and regulators of cell growth and division, including many genes with unknown functions were uncovered. A Northern blot analysis was used to verify the expression pattern of the corresponding genes under metal treatment. The metal ions enhanced the expression of alcohol dehydrogenase Adh1 homologue by up to 33-fold in A. flavus and ca fourfold in F. graminearum. Encoding homologues of a neutral amino acid permease, were also used in the Northern analysis. However, the expression of the permease was not significantly affected by metal ion treatments. CONCLUSIONS: The results showed a significant effect of metal ions on expression of gene related to fungal growth, development, conidiation and production of both aflatoxin and zearalenone. SIGNIFICANT AND IMPACT OF THE STUDY: At the molecular and cellular level, the significant effects of metal ions on fungal growth and development, conidiation, and production of both aflatoxin and zearalenone were demonstrated.

Aflatoxins↗

PubMed2006

Binding of Fusarium mycotoxins by fermentative bacteria in vitro.

AIMS: Fusarium toxins can occur in conserved forages impairing farm animal performances and health. On-farm biological decontamination methods could be an alternative to traditional physico-chemical methods. In this work, the ability to remove Fusarium toxins by fermentative bacteria was evaluated in vitro. METHODS AND RESULTS: Twenty-nine strains of lactic (LAB) and propionic acid bacteria (PAB) were tested for their ability to remove deoxynivalenol (DON) and fumonisins B1 and B2 (FB1, FB2) from an acid, pH 4, medium. Mycotoxin removal was widespread for LAB, but differences among strains were large. Removal was up to 55% for DON, 82% for FB1 and 100% for FB2. Selected strains were also capable of removing up to 88% zearalenone. The PAB strains were less efficient than the LAB. Binding, not biodegradation appeared to be the mode of action, as no toxin derivatives were observed and removal was not impaired in nonviable bacteria. Binding was not affected by pH, except for fumonisins that decreased to nearly 0% at neutral pH. CONCLUSIONS: Selected fermentative bacteria are able to bind main Fusarium mycotoxins. SIGNIFICANCE AND IMPACT OF THE STUDY: The binding ability of selected strains could be used to decrease the bioavailability of toxins in contaminated silages.

Culture Media↗

PubMedDate not supplied

Effects of Aspergillus sulphureus mycotoxins on Cryptococcus neoformans.

Many studies have evaluated the toxicity of mycotoxins to mammals, but there is little information on their action against fungal cells, even although mycotoxins are frequently active against fungi in nature. A crude extract of Aspergillus sulphureus was tested for its growth-inhibitory effect on Cryptococcus neoformans. The reduction in cell growth of Cr. neoformans caused by the extract was dose dependent. Using a liquid medium containing 2% A. sulphureus extract, the RNA content of Cryptococcus amounted to about 60% of that of non-treated cells. Capsule thickening, demonstrated biochemically and with cytological stains, occurred at doses that had minimal effect on cell growth and RNA content. Our results suggest that the virulence of Cr. neoformans may increase in cases of coenobiosis with A. sulphureus, which is theoretically possible in places where corn-fed pigeons are numerous.

Aspergillus↗

PubMed1990

The use of tissue culture for the detection of mycotoxins.

After incubation with 20 different mycotoxin standards and extracts from fungi and feed stuffs, fluorescent flow cytometry was used to measure viability of NS-1 cells and compared to microscopic assessment of cytotoxicity on stained HEp-11 monolayers. Both methods gave essentially the same results but the cytometric analysis offered a more quantitative approach and was particularly appropriate in the screening of extracts containing fats. The potential uses of a cytotoxic test in the analysis of feed stuffs and fungal extracts for the presence of mycotoxins are discussed.

Animals↗

PubMed2006

Influence of water activity, temperature and time on mycotoxins production on barley rootlets.

AIMS: The objective of this study was to determine the ochratoxin (OT) and aflatoxin (AF) production by three strains of Aspergillus spp. under different water activities, temperature and incubation time on barley rootlets (BR). METHODS AND RESULTS: Aspergillus ochraceus and Aspergillus flavus were able to produce mycotoxins on BR. Aspergillus ochraceus produced ochratoxin A (OTA) at 0.80 water activity (a(w)), at 25 and 30 degrees C as optimal environmental conditions. The OTA production varies at different incubation days depending on a(w). Aflatoxin B(1) (AFB1) accumulation was obtained at 25 degrees C, at 0.80 and 0.95 a(w), after 14 and 21 incubation days respectively. Temperature was a critical factor influencing OTA and AFB(1) production. CONCLUSIONS: This study demonstrates that BR support OTA and AFB(1) production at relatively low water activity (0.80 a(w)) and high temperatures (25-30 degrees C). SIGNIFICANCE AND IMPACT OF THE STUDY: The study of ecophysiological parameters and their interactions would determine the prevailing environmental factors, which enhance the mycotoxin production on BR used as animal feed.

Aflatoxins↗

PubMed2006

Role of zearalenone lactonase in protection of Gliocladium roseum from fungitoxic effects of the mycotoxin zearalenone.

Zearalenone is a mycotoxin with estrogenic effects on mammals that is produced by several species of Fusarium. We found that zearalenone and its derivatives inhibit the growth of filamentous fungi on solid media at concentrations of < or =10 microg/ml. The fungitoxic effect declined in the order zearalenone > alpha-zearalenol > beta-zearalenol. The mycoparasitic fungus Gliocladium roseum produces a zearalenone-specific lactonase which catalyzes the hydrolysis of zearalenone, followed by a spontaneous decarboxylation. The growth of G. roseum was not inhibited by zearalenone, and the lactonase may protect G. roseum from the toxic effects of this mycotoxin. We inactivated zes2, the gene encoding zearalenone lactonase in G. roseum, by inserting a hygromycin resistance cassette into the coding sequence of the gene by means of Agrobacterium tumefaciens-mediated genetic transformation. The zes2 disruption mutants could not hydrolyze the lactone bond of zearalenone and were more sensitive to zearalenone. These data are consistent with a hypothesis that resorcylic acid lactones exemplified by zearalenone act to reduce growth competition by preventing competing fungi from colonizing substrates occupied by zearalenone producers and suggest that they may play a role in fungal defense against mycoparasites.

Carboxylic Ester Hydrolases↗

PubMed1979

Blue light inhibits mycotoxin production and increases total lipids and pigmentation in Alternaria alternata.

Light inhibits production of the mycotoxins alternariol and alternariol monomethyl ether, both polyketids produced by Alternaria alternata. This effect seems to be general because seven isolates of A. alternata with different alternariol- and alternariol monomethyl ether-producing abilities all respond to continuous light with reduced levels of alternariol and alternariol monomethyl ether when the mycotoxins were calculated on a microgram-per-milligram (dry weight) basis. Blue light inhibited alternariol and alternariol monomethyl ether production 69 and 77%, respectively. Red light gave no reduction of toxin levels. Total lipids were increased 25% when mycelium was grown in blue light as compared with red light or darkness. In white or blue light, but not in red light or darkness, a red-brown pigment accumulated by the mycelium.

Carotenoids↗

PubMed1983

Comparison of cytotoxicity and thin-layer chromatography methods for detection of mycotoxins.

Thirty-three standard mycotoxins were assayed by thin-layer chromatography and by cytotoxicity in HEp-2 and Chang cells. Various levels of detection were found. The cytotoxicity test was significantly more sensitive than thin-layer chromatography for the trichothecenes and should be useful for screening extracts from animal feedstuffs for the presence of unknown mycotoxins.

Biological Assay↗

PubMed1984

Effect of water activity and temperature on mycotoxin production by Alternaria alternata in culture and on wheat grain.

Both water activity (aW) and temperature affected the production of altenuene (AE), alternariol (AOH), and alternariol monomethyl ether (AME) by Alternaria alternata on wheat extract agar and wheat grain. Greatest production of all three mycotoxins occurred at 0.98 aW and 25 degrees C on both substrates. At 0.98 aW and 25 degrees C, a single colony of A. alternata grown on wheat extract agar produced 807 micrograms of AOH, 603 micrograms of AME, and 169 micrograms of AE ml in 30 days. However, production of all three mycotoxins at 0.95 aW was less than 40% of these amounts. Little toxin was produced at 0.90 aW. Changing temperature and aW altered the relative amounts of the different toxins produced on agar. At 15 degrees C and 0.98 aW, maxima of 52 micrograms of AOH and 25 micrograms of AME per ml were produced after 15 and 30 days, respectively, whereas AE continued to increase and reached 57 micrograms/ml after 40 days. At 15 degrees C and 0.95 aW, production was, respectively, 62, 10, and 5 micrograms/ml after 40 days. All three metabolites were produced at 5 degrees C and 0.98 to 0.95 aW and at 30 degrees C and 0.98 to 0.90 aW. On wheat grain at 25 degrees C and 0.98 to 0.95 aW, more AME was produced than AOH or AE, but at 15 degrees C there was less AME than AOH or AE. Only trace amounts of AE, AOH, and AME were found at 15 to 25 degrees C and 0.90 aW, but production of AME was inhibited at 30 degrees C and 0.95 aW or less.

Alternaria↗

PubMed1986

Mycological survey of Korean cereals and production of mycotoxins by Fusarium isolates.

The fungal species isolated from Korean cereals (barley, polished barley, wheat, rye, and malt) were Alternaria spp., Aspergillus spp., Chaetomium spp., Drechslera spp., Epicoccum sp., Fusarium spp., and Penicillium spp., etc. The number of Fusarium strains isolated was 36, and their ability to produce Fusarium mycotoxins on rice was tested. Nivalenol (NIV) was produced by Fusarium graminearum (7 of 9 isolates), Fusarium oxysporum (3 of 10 isolates), and Fusarium spp. (7 of 15 isolates). Of 15 isolates of Fusarium spp., 6 formed deoxynivalenol (DON). Fusarenon-X and 3-acetyl-DON were produced by most NIV- and DON-forming isolates, respectively. Zearalenone was produced by 3 isolates of F. graminearum, 1 isolate of Fusarium equiseti, and 11 isolates of Fusarium spp. T-2 toxin was not produced by any Fusarium isolates. The highest concentrations of mycotoxins produced by Fusarium isolates were 77.4 (NIV), 5.3 (DON), 138.3 (fusarenon-X), 40.6 (3-acetyl-DON), and 23.2 (zearalenone) micrograms/g.

Chaetomium↗

PubMed1987

Tremorgenic mycotoxins from Aspergillus fumigatus as a possible occupational health problem in sawmills.

Wood-trimmers' disease, generally called extrinsic allergic alveolitis, which affects workers in sawmills, is thought to be caused by fungal diaspores. The importance of Aspergillus fumigatus on the surface of wood dried in kilns is accentuated by its ability to produce tremorgenic mycotoxins. Eight strains of A. fumigatus from five different sawmills were isolated and cultivated on liquid media, and one of the strains was also cultivated on wood blocks. Extracts were prepared, and the tremorgenic reactions were induced by oral administration of extracts to rats. Extracts of the strain grown in liquid medium and on wood blocks induced very strong tremorgenic reactions when administered orally to rats. Four other strains induced mild tremorgenic reactions. High-performance liquid chromatography analysis revealed two tremorgenic mycotoxins, verruculogen and fumitremorgen C, in the five toxic strains. One nontoxic strain produced detectable levels of verruculogen. These results, coupled with the known resemblance of the acutely toxic phase of wood-trimmers' disease to the symptoms produced by these tremorgens, imply that wood-trimmers' disease and similar occupational diseases are, at least in part, mycotoxicoses.

Air Microbiology↗

PubMed1996

Natural occurrence of Fusarium mycotoxins in field samples from the 1992 Wisconsin corn crop.

Analysis of 98 moldy corn samples collected in Wisconsin between November 1992 and January 1993 for Fusarium toxins by various immunochemical assays revealed overall average mycotoxin concentrations of 305.6, 237.7, and 904.3 ng/g for type A trichothecenes (TCTCs), deoxynivalenol (DON)-related type B TCTCs (total DON), and zearalenone (ZE), respectively. A small portion (5.1%) of the samples was found to be contaminated with high levels ( > 1 microgram/g) of type A TCTCs and total DON during the whole survey. Over 40% of the samples had 100 to 1,000 ng of total DON per g, while 17% of the samples had the same levels of type A TCTCs. The analytical data were consistent with those from mycological examinations for the samples in which various toxic Fusarium spp., including F. sporotrichioides, F. poae, and F. graminearum, were found. The samples received in November 1992 had relatively low concentrations of toxin; the average levels of type A TCTCs and total DON were 9.9 and 79 ng/g, respectively. The toxin concentrations became progressively higher in the samples received in December. The average levels for the type A TCTCs and total DON increased to 920 and 335 ng/g, respectively. However, the levels of ZE were higher in the samples collected earlier. The average levels for samples collected in November and late December were 1,195 and 242 ng/g, respectively. Analysis of selected samples by high-performance liquid chromatography monitoring with an enzyme-linked immunosorbent assay revealed that T-2 toxin, HT-2 toxin, diacetoxyscirpenol, neosolaniol, and T-2 tetraol (T-2-4ol) were common in these samples. Statistical analysis revealed a weak correlation between the levels of total type A TCTCs and total DON in the samples (r = 0.18, P = 0.09), but a strong correlation between the levels of ZE and total type B TCTCs (r = 0.75, P < 0.0001) was found. The mycotoxin levels of total type A TCTCs, total DON-related type B TCTCs, and ZE in the cobs (5.2, 3.9, and 21 micrograms/g, respectively) were considerably higher than those in the kernels (1.0, 0.5, and 0.5 microgram/g, respectively). The type A toxin levels increased from a range of 14 to 35 ng/g to a range of 110 to 538 ng/g after the moldy corn samples were held at 5 degrees C for 8 days in the laboratory.

Food Microbiology↗

PubMed2003

Mycotoxin fumonisin B1 increases intestinal colonization by pathogenic Escherichia coli in pigs.

Fumonisin B(1) (FB(1)) is a mycotoxin that commonly occurs in maize. FB(1) causes a variety of toxic effects in different animal species and has been implicated as a contributing factor of esophageal cancers in humans. In the present study, we examined the effect of dietary exposure to FB(1) on intestinal colonization by pathogenic Escherichia coli associated with extraintestinal infection. Three-week-old weaned pigs were given FB(1) by gavage as a crude extract or as a purified toxin at a dose of 0.5 mg/kg of body weight daily for 6 days. On the last day of the toxin treatment, the pigs were orally inoculated with an extraintestinal pathogenic E. coli strain. All animals were euthanized 24 h later, necropsies were performed, and tissues were taken for bacterial counts and light microscopic examination. Ingestion of FB(1) had only a minimal effect on animal weight gain, did not cause any macroscopic or microscopic lesions, and did not change the plasma biochemical profile. However, colonization of the small and large intestines by an extraintestinal pathogenic E. coli strain was significantly increased. Our results show that FB(1) is a predisposing factor to infectious disease and that the pig can be used as a model for the study of the consequences of ingesting mycotoxin-contaminated food.

Administration, Oral↗

PubMed2005

Detection of airborne Stachybotrys chartarum macrocyclic trichothecene mycotoxins in the indoor environment.

The existence of airborne mycotoxins in mold-contaminated buildings has long been hypothesized to be a potential occupant health risk. However, little work has been done to demonstrate the presence of these compounds in such environments. The presence of airborne macrocyclic trichothecene mycotoxins in indoor environments with known Stachybotrys chartarum contamination was therefore investigated. In seven buildings, air was collected using a high-volume liquid impaction bioaerosol sampler (SpinCon PAS 450-10) under static or disturbed conditions. An additional building was sampled using an Andersen GPS-1 PUF sampler modified to separate and collect particulates smaller than conidia. Four control buildings (i.e., no detectable S. chartarum growth or history of water damage) and outdoor air were also tested. Samples were analyzed using a macrocyclic trichothecene-specific enzyme-linked immunosorbent assay (ELISA). ELISA specificity was tested using phosphate-buffered saline extracts of the fungal genera Aspergillus, Chaetomium, Cladosporium, Fusarium, Memnoniella, Penicillium, Rhizopus, and Trichoderma, five Stachybotrys strains, and the indoor air allergens Can f 1, Der p 1, and Fel d 1. For test buildings, the results showed that detectable toxin concentrations increased with the sampling time and short periods of air disturbance. Trichothecene values ranged from <10 to >1,300 pg/m3 of sampled air. The control environments demonstrated statistically significantly (P < 0.001) lower levels of airborne trichothecenes. ELISA specificity experiments demonstrated a high specificity for the trichothecene-producing strain of S. chartarum. Our data indicate that airborne macrocyclic trichothecenes can exist in Stachybotrys-contaminated buildings, and this should be taken into consideration in future indoor air quality investigations.

Air Microbiology↗

PubMed2000

Effects of the Fusarium spp. mycotoxins fusaric acid and deoxynivalenol on the growth of Ruminococcus albus and Methanobrevibacter ruminantium.

The Fusarium spp. mycotoxins fusaric acid and deoxynivalenol (DON) were tested for antimicrobial activity against Ruminococcus albus and Methanobrevibacter ruminantium. The growth of both organisms was inhibited by fusaric acid as low as 15 micrograms/mL (84 microM) but not by DON, at levels as high as 100 micrograms/mL (338 microM). No synergistic inhibitory effect was observed with DON plus fusaric acid. Neither organism was able to adapt to the fusaric acid and responses of each organism to the compound were different. The optical density (OD) maximum for R. albus, but not for M. ruminantium, was diminished after 28 days incubation at concentrations of fusaric acid below 240 micrograms/mL. Inhibition of R. albus started before significant growth had occurred, while M. ruminantium doubled twice before the onset of inhibition. Responses to picolinic acid, an analog of fusaric acid, were also dramatically different between the two microorganisms with M. ruminantium exhibiting a severe lag followed by a complete recovery of growth, while R. albus was only slightly inhibited with no lag. These results suggest that the mechanism of fusaric acid inhibition is specific to each microorganism. This is the first demonstration of the common mycotoxin fusaric acid inhibiting the growth of rumen bacteria.

Animals↗