Dexamethasone treatment of lipopolysaccharide-induced meningitis in rabbits that mimics magnification of inflammation following antibiotic therapy

Natural occurrence of fumonisins B1 and B2, incidence of Fusarium species, and capacity to produce fumonisins by Fusarium isolates, were investigated in 50 corn-based samples from Spain destined for animal consumption. Forty-four samples (88%) were found to be contaminated with fumonisins. The levels of contamination were very low, with a mean of 400 ng/g in the samples. We investigated the capacity of 11 isolates of Fusarium moniliforme and 19 isolates of F. proliferatum to produce fumonisins. All F. proliferatum isolates and 8 out of the 11 F. moniliforme isolates assayed produced fumonisins on a corn medium. The FB1/FB2 ratio in the isolates ranged from 1.1 to 3.5.     

Measurement of antibacterial activities of T-2 toxin, deoxynivalenol, ochratoxin A, aflatoxin B1 and fumonisin B1 using microtitration tray-based turbidimetric techniques

Various mycotoxins were tested for their antibacterial activity by evaluating growth delays using a fully automated microturbidmetric method. Ten different strains of the genera Escherichia, Streptococcus, Staphylococcus, Yersinia, Salmonella, Erysipelothrix and Lactobacillus were used as test micro-organisms. T-2 toxin, deoxynivalenol (DON), ochratoxin A (OTA), aflatoxin B1 (AFB1) and fumonisin B1 (FB1) were used as representative mycotoxins. The inhibitory effect in vitro was defined as the difference between the growth rate without mycotoxins and the growth rate in the presence of a mycotoxin. Among the tested strains, Streptococcus agalactiae was found to be sensitive to all the toxins, with the exception of OTA. T-2 toxin and FB1 were the most effective in slowing down the growth of Staphylococcus aureus. AFB1 affected the growth of Yersinia enterocolitica. The growth rate of Escherichia coli and Salmonella infantis was decreased by FB1. Among the bacterial strains used in this study, only the growth of Erysipelothrix rhusiopathiae was inhibited by OTA. Thus, using appropriate tester strains it should be possible to set up a broad-range microtubidimetry assay for individual mycotoxin screening in vitro. We concluded that the microtitration technique provides a rapid, convenient and high-throughput capacity system to analyse bacteria-mycotoxin interactions.     

Total hip arthroplasty cemented femoral component distal stem centralizer. Effect on stem centralization and cement mantle

Samples of wheat harvested from 1988 to 1990 and stored in elevators in the south of Brazil (12 Brazilian, 4 Argentinian and 2 Uruguayan) were analysed in 1990 for 14 mycotoxins: deoxynivalenol (DON), nivalenol, diacetoxyscirpenol (DAS), T-2 and HT-2 toxins, T-2 triol, T-2 tetraol, aflatoxins B1, B2, G1, G2, ochratoxin A (OCHRA A), zearalenone and sterigmatocystin. One sample (1988 harvest) was contaminated with OCHRA A (0.04 microgram/g) and three other samples (1990 harvest) were contaminated with DON (0.40 microgram/g), DAS (0.30 microgram/g), T-2 (two samples, 0.35 and 0.36 gamma g/g) and T-2 tetraol (1.68 micrograms/g). Fusarium graminearum Schwabe was found in the 1990 samples with a relative incidence ranging from 1 to 22% and predominated in Argentinian and Uruguayan wheat (1990 harvest). Fusarium dimerum Penzig (8-75%) was the main Fusarium sp. in Brazilian wheat from the 1990 harvest.     

Stability of fumonisins in thermally processed corn products

Little is known about the stability of fumonisins in corn-based foods during heating. This study investigated the effects of canning, baking, and roasting (dry heating) processes on the stability of fumonisins in artificially contaminated and naturally contaminated corn-based foods. All samples were analyzed for fumonisin levels by both a commercial enzyme-linked immunosorbent assay (ELISA) and a high-performance liquid chromatographic (HPLC) method. Canned whole-kernel corn showed a significant (P < or = 0.05) decrease in fumonisins by both ELISA (15%) and HPLC (11%) analyses. Canned cream-style corn and baked corn bread showed significant (P < or = 0.05) decreases in fumonisin levels at an average rate of 9% and 48%, respectively, as analyzed by ELISA. Corn-muffin mix artificially contaminated with 5 micrograms of fumonisin B1 (FB1) per g and naturally contaminated corn-muffin mix showed no significant (P < or = 0.05) losses of fumonisins upon baking. Roasting cornmeal samples artificially contaminated with 5 micrograms of FB1 per g and naturally contaminated cornmeal samples at 218 degrees C for 15 min resulted in almost complete loss of fumonisins.     

Characterization of cell-cycle arrest by fumonisin B1 in CV-1 cells

Fusarium moniliforme is a widespread fungal pathogen which primarily infects corn, but can also infect rice or wheat. Fusarium moniliforme produce several mycotoxins, the most prominent of which is called fumonisin B1 (FB1). Epidemiological studies have indicated that ingestion of fumonisins correlates with a higher incidence of oesophageal cancer in Africa and China. Fumonisins also cause a neurodegenerative disease in horses, induce hepatic cancer in rats, are nephrotoxic in rats, or cause pulmonary oedema in swine. Structurally, fumonisins resemble sphingolipids and can alter sphingolipid biosynthesis. suggesting that sphingolipid alterations play a role in disease and carcinogenesis. Previous studies determined that FB1 blocked cell-cycle progression in CV-1 cells but not COS-7 cells. Herein, we have examined the effects that FB1 treatment has on cell-cycle regulatory proteins. Our studies established that FB1 treatment of CV-1 cells, but not COS-7 cells, leads to dephosphorylation of the retinoblastoma (Rb) protein. Cyclin dependent kinase 2 (CDK2) activity was repressed five- to 10-fold and cyclin E protein levels were lower in CV-1 cells after fumonisin treatment. Two CDK inhibitors, Kip1 and Kip2, were induced within 3 hours after fumonisin treatment of CV-1 cells, suggesting these two proteins mediate cell-cycle arrest induced by FB1. This mycotoxin caused large increases in sphinganine within 3 hours after addition of FB1. As sphingoid bases are known to induce Rb phosphorylation, this increase in sphinganinie might be the stimulus for the suppression of cyclin dependent kinase activities via Kip1 and Kip2. The ability of FB1 to accumulate sphingosine or sphinganine and arrest the cell cycle in some cells but not others may play an important role in carcinogenesis or disease.     

Schistosomiasis vaccine development--the current picture

The fate of three Fusarium mycotoxins, nivalenol (NIV), deoxynivalenol (DON) and zearalenone (ZEN), all common contaminants in New Zealand-grown maize, has been measured in fractions of maize after passage through a commercial wet-milling plant. Distribution of the three toxins follows a pattern reasonably expected from their physical solubility characteristics. The highly water-soluble mycotoxins, NIV and DON, were found at high concentrations (up to 8.8 mg/kg) in concentrated steep liquor (CSL) fractions, but at low levels (less than 0.3 mg/kg) in the solid (germ, fibre and gluten) fractions. The converse was true for ZEN, which is relatively insoluble in water. For ZEN, the maximum concentration found in CSL was 0.6 mg/kg compared with 2.2-4.8 mg/kg in germ, fibre and gluten fractions. Accordingly, an animal food byproduct composed mainly of pressed fibre and concentrated steep liquor was usually found to contain concentrations of all three mycotoxins above those existing in the input maize. A single sample of corn oil recovered during the study also had a high concentration (4.6 mg/kg) of ZEN. The analytical clean-up method used converts all trichothecenes present to parent alcohols, therefore results are indicative of total trichothecene content. HPLC analytical conditions suitable for the analysis of NIV and DON in complex process grain products are also described.     

Occurrence of fumonisins in corn-based food products

Corn-based food products obtained from commercial outlets in three different parts of the U.S., Maryland, Nebraska, and Arizona were analyzed for total fumonisins by a commercial competitive direct enzyme-linked immunosorbent assay (CD-ELISA) and for fumonisin B1 (FB1) by high-performance liquid chromatography (HPLC). The highest fumonisin concentrations were found in samples collected in Maryland, where all 18 samples were found positive for fumonisins (200 to 7,450 ng/g of food) by CD-ELISA and 15 of the 18 samples (83%) were found positive for FB1 (< 75 to 5,916 ng/g) by HPLC. Fumonisins were also detected by CD-ELISA in 14 of 15 samples collected in Arizona with concentrations ranging from 200 to 1,450 ng/g, but analyses by HPLC showed that only 8 of 15 samples (53%) were positive for FB1 (< 75 to 1,565 ng/g of food). Of the 23 samples collected in Nebraska, 20 (87%) were positive for fumonisins (200 to 2,500 ng/g) by CD-ELISA, but only 10 (44%) were positive for FB1 (< 75 to 927 ng/g) by HPLC. The highest fumonisin and FB1 concentrations were found in cornmeal samples, ranging up to 7,450 ng/g of cornmeal by CD-ELISA and 5,916 ng/g by HPLC. These findings indicate that there may be a risk of human exposure to fumonisins through the consumption of some corn-based foods.     

Quantitative glycohistochemistry defines new prognostic markers for cancers of the oral cavity

Fumonisin B1 (FB1) and aminopentol (AP1) (which is formed by hydrolysis of FB1) are found in corn contaminated with some strains of Fusarium moniliforme. Incubation of HT29 cells (a human colonic cell line) with FB1 or AP1 caused a significant reduction in cell number; AP1 was less potent, with 50 microM AP1 causing the same reduction (ca. 30% after 24 h) as 10 microM FB1. The reduction in cell number reflected increases in DNA fragmentation and the percentage of apoptotic cells. Both FB1 and AP1 caused the accumulation of sphinganine (25- and 35-fold by 10 microM FB1 and 50 microM AP1, respectively); thus, concentrations of FB1 and AP1 that caused comparable reductions in cell number were also similar with respect to elevation of sphinganine, a compound that is growth inhibitory and cytotoxic. Inhibition of the first step of sphingolipid biosynthesis with ISP-1 prevented the elevation in sphinganine, DNA fragmentation, and apoptosis induced by FB1. Therefore, these effects of FB1 on HT29 cells can be attributed to the accumulation of sphinganine. Since consumption of food contaminated with Fusarium moniliforme (Sheldon) exposes colonic cells to these mycotoxins, the possibility that FB1 and AP1 are toxic for intestinal cells in vivo should be evaluated, especially in the light of the recent report (Bhat et al., Clin. Toxicol. 35, 249, 1997) describing intestinal disturbances in humans after consumption of moldy corn and sorghum containing fumonisins.     

Studies of 99mTc-BnAO (HL-91): a non-nitroaromatic compound for hypoxic cell detection

Fumonisins are toxic metabolites of Fusarium moniliforme, a fungus that occurs widely in corn. Fumonisins causes leukoencephalomalacia in horses and pulmonary edema in swine and have been suggested as a possible cause of an increased incidence of neural tube defects among people living along the Texas-Mexico border. As part of an effort to determine levels of fumonisins in human food, a liquid chromatographic (LC) method was devised for determining fumonisin B1 (FB1) and the total hydrolysis product of FB1 (HB1) in tortillas. The method uses acetonitrile-0.1M phosphate buffer (pH 3; 1 + 1) extraction, solid-phase C18 cleanup, o-phthalaldehyde and 2-mercaptoethanol derivatization, and reversed-phase LC. Average recoveries from tortillas spiked with FB1 and HB1 at 250, 500, and 1000 ng/g were 86.5% for FB1 and 82.6% for HB1. Tortillas (54) and masa (8) from the Texas-Mexico border were analyzed for FB1 and HB1. Average amounts of FB1 and HB1 in tortillas were 187 and 82 ng/g, respectively. Average amounts of FB1 and HB1 in masas were 262 and 64 ng/g, respectively. The results show that fumonisin B1 and its hydrolysis product are present in tortillas consumed by a population experiencing an increased incidence of neural tube defects.     

Aflatoxicosis in turkey poults is prevented by treatment of naturally contaminated corn with ozone generated by electrolysis

Previous studies have demonstrated that a novel source of ozone gas (O3) maybe used to chemically degrade numerous mycotoxins, including aflatoxin (AF) B1. Subsequent in vitro analyses demonstrated detoxification of AFB1, suggesting a potential method of remediate AF-contaminated grain. The objective of this study was to evaluate the capability of electrochemically produced ozone to degrade AFB1 in naturally contaminated whole kernel corn and confirm detoxification in turkey poults. Corn was procured from the southern coastal areas of Texas and HPLC revealed 1,220 +/- 73.3 ppb AFB1. Control and contaminated corn were treated for 92 h with O3 at 200 mg/min in 30 kg batches; greater than 95% reduction of AFB1 in contaminated corn was achieved. One-day-old female turkey poults were fed 1) control corn, 2) control corn + O3, 3) AFB1 corn, or 4) AFB1 corn + O3 mixed in rations (46% by wt.) and consumed ad libitum for 3 wk. When compared with controls, turkeys fed AFB1 corn had reduced body weight gain and relative liver weight, whereas turkeys fed control corn + O3 or AFB1 corn + O3 did not differ from controls. Furthermore, alterations in the majority of relative organ weight, liver discoloration, serum enzyme activity, hematological parameters, and blood chemistry caused by AFB1 were eliminated (no difference from controls) by treatment with O3. These data demonstrate that treatment of contaminated corn with electrochemically produced O3 provided protection against AFB1 in young turkey poults. It is important to note that treatment of control corn with O3 did not alter the performance of the turkey poults.     

Effect of cimetidine on basal and postprandial plasma concentrations of cholecystokinin and gastrin in humans

Strains (10705) of microscopic fungi were isolated from spring barley heads in the region of Lublin (south-eastern Poland). Fusarium sporotrichioides Sherb was found in 418 (3.9%) of isolated strains. Group A trichothecene mycotoxins were detected in the collected barley kernels colonized by F. sporotrichioides, with Fusarium head blight symptoms. Among 24 samples analysed, 12 were T-2 toxin positive in a range of contamination from 0.02 to 2.40 micrograms/g (average 0.45), while in five samples HT-2 toxin ranged from 0.01 to 0.37 micrograms/g (average 0.23) and T-2 tetraol was detected in two samples in a range of 0.01-0.21 micrograms/g (average 0.11). Twelve samples were free of detectable amounts of the toxic metabolites analysed.     

The effect of traumatic brain injury on children with learning disability

In Argentina, there is rather little information about the natural occurrence of mycotoxins in feedstuffs. The aim of this work was to determine the fungal flora and natural incidence of aflatoxin B1 (AFB1), zearalenone (ZEA) and deoxynivalenol (DON) in poultry feeds from 5 factories of Río Cuarto, Córdoba. Three hundred samples were taken from May 1995 to May 1996. Fungal counts of poultry feeds ranged 10(4) to 10(6) CFU g-1. The lowest counts were obtained on the first months from the sampling (May to September 1995) with mean values significantly different from those found at the last of the sampling (October 1995 to April 1996). The most prevalent species isolated of poultry feed samples belonged to the genera Penicillium that was present in 98% of the samples, Fusarium (87%) and Aspergillus (52%). Fusarium species isolated were: F moniliforme in 73% of the samples, F subglutinans (35%), F graminearum (20%) and within Aspergillus species: A. parasiticus (33%) and A. flavus (8%) were identified. In poultry feeds aflatoxin B1 (AFB1) was the most significant mycotoxin with levels ranging from 17 to 197 ng/g. For deoxynivalenol (DON) the levels ranged from 240 to 410 ng/g. Only three out of 300 samples were contaminated with zearalenone (ZEA) in concentrations of 30, 120 and 280 ng/g. These are preliminary data on this subject in our region.     

Acylation of naturally occurring and synthetic 1-deoxysphinganines by ceramide synthase. Formation of N-palmitoyl-aminopentol produces a toxic metabolite of hydrolyzed fumonisin, AP1, and a new category of ceramide synthase inhibitor

Fumonisin B1 (FB1) is the predominant member of a family of mycotoxins produced by Fusarium moniliforme (Sheldon) and related fungi. Certain foods also contain the aminopentol backbone (AP1) that is formed upon base hydrolysis of the ester-linked tricarballylic acids of FB1. Both FB1 and, to a lesser extent, AP1 inhibit ceramide synthase due to structural similarities between fumonisins (as 1-deoxy-analogs of sphinganine) and sphingoid bases. To explore these structure-function relationships further, erythro- and threo-2-amino, 3-hydroxy- (and 3, 5-dihydroxy-) octadecanes were prepared by highly stereoselective syntheses. All of these analogs inhibit the acylation of sphingoid bases by ceramide synthase, and are themselves acylated with Vmax/Km of 40-125 for the erythro-isomers (compared with approximately 250 for D-erythro-sphinganine) and 4-6 for the threo-isomers. Ceramide synthase also acylates AP1 (but not FB1, under the conditions tested) to N-palmitoyl-AP1 (PAP1) with a Vmax/Km of approximately 1. The toxicity of PAP1 was evaluated using HT29 cells, a human colonic cell line. PAP1 was at least 10 times more toxic than FB1 or AP1 and caused sphinganine accumulation as an inhibitor of ceramide synthase. These studies demonstrate that: the 1-hydroxyl group is not required for sphingoid bases to be acylated; both erythro- and threo-isomers are acylated with the highest apparent Vmax/Km for the erythro-analogs; and AP1 is acylated to PAP1, a new category of ceramide synthase inhibitor as well as a toxic metabolite that may play a role in the diseases caused by fumonisins.     

The socioeconomic impact of chronic back pain: is anyone benefiting?

The trichothecene 3-O-acetyltransferase gene, Tri101, plays a pivotal role for the well-being of the type B trichothecene producer Fusarium graminearum. We have analyzed the cosmids containing Tri101 and found that this resistance gene is not in the biosynthetic gene cluster reported so far. It was located between the UTP-ammonia ligase gene and the phosphate permease gene which are not related to trichothecene biosynthesis. These two 'house-keeping' genes were also linked in Fusarium species that do not produce trichothecenes. The result suggests that the isolated occurrence of Tri101 is attributed to horizontal gene transfer and not to the reciprocal translocation of the chromosome containing the gene cluster. Interestingly, 3-O-acetylation was not always a primary self-defensive strategy for all the t-type trichothecene producers; i.e. the type A trichothecene producer Fusarium sporotrichioides did not acetylate T-2 toxin in vivo although the fungus possessed a functional 3-O-acetyltransferase gene. Thus Tri101 appears to be a defense option which the producers have independently acquired in addition to their original resistance mechanisms.     

Analysis of fumonisins and Alternaria alternata toxin by liquid chromatography-enzyme-linked immunosorbent assay

Use of a direct competitive enzyme-linked immunosorbent assay (ELISA) as a postcolumn monitoring system after liquid chromatography (LC) is described for analysis of different fumonisin analogs. Without cleanup and derivatization, sample extracts are directly injected into a C18 reversed-phase column and then subjected to LC. Fractions (0.5 mL each) are collected and then analyzed by ELISA. LC using a water-methanol gradient separated the 3 major fumonisins FmB1, FmB2, and FmB3, and as low as 0.1 ng FmB1 could be detected. Recovery of FmB1 added to ground corn (100-1000 ng/g) and then extracted with CH3CN-H2O (1 + 1, v/v) was 78.8%. Analysis of fumonisins in one starch and 14 naturally contaminated corn samples showed that FmB1 was the major fumonisin. Ten samples also were contaminated with FmB2, but only 2 samples were contaminated with FmB3. The method also was used to analyze extracts from cultures of 3 Alternaria alternata (AAL) strains. Both FmB1 and the AAL toxin TA were detected in the culture extracts, and their amounts varied considerably with the cultures tested.     

Influence of fumonisin B1, present in Fusarium moniliforme culture material, and T-2 toxin on turkey poults

Diets containing 300 mg fumonisin B1 (FB1)/kg of feed and 5 mg T-2 toxin/kg of feed singly or in combination were fed to female turkey poults (Nicholas Large White) from day of hatch to 21 d of age. When compared with controls, 21-d body weight gains were reduced 21% by FB1, 26% by T-2, and 47% by the combination. the efficiency of feed utilization was adversely affected by FB1 and the combination of FB1 and T-2. Relative weights (grams/100 g BW) of the liver and gizzard were increased in poults fed the FB1 and the combination diets; whereas, the relative weight of the pancreas was increased in all treated groups. All poults were scored for oral lesions using a scale of 1 to 4 (1 = no visible lesions, 4 = severe lesions). Oral lesions were present in all poults fed the T-2 diet (average score of 3.29) or the combination diet (average score of 3.54). Serum concentration of cholesterol was decreased and lactate dehydrogenase activity was increased in poults fed the FB1 and combination diets. The activity of aspartate aminotransferase and the values for red blood cells, hemoglobin, and hematocrit were increased only in poults fed the combination diet. Inorganic phosphorus concentration was decreased only in poults fed the combination diet. The increased toxicity in poults fed the combination diet for most variables can best be described as additive, although some variables not altered by FB1 or T-2 singly were significantly affected by the combination, indicating that the combination may pose a potentially greater problem to the turkey industry than either of the mycotoxins individually.     

Dietary fumonisins disrupt sphingolipid metabolism in mink and increase the free sphinganine to sphingosine ratio in urine but not in hair

This study was conducted to investigate the effects of dietary Fusarium moniliforme culture material (M-1325) containing known concentrations of fumonisins B1, B2 and B3 on sphingolipids in urine and hair of mink (Mustela vison) for use as potential, non-invasive biomarkers of exposure to fumonisins in this species. Feeding mink diets containing 86, 22, and 7 ppm or 200, 42, and 12 ppm of fumonisins B1, B2 and B3, respectively, yielded marked increases in urinary free sphinganine (Sa) and free sphingosine (So) concentrations, and free Sa/free So ratios (2 to 11-fold) within 7 d, compared to controls. Free Sa and free So concentrations and Sa/So ratios in hair samples from mink fed the control or high dose fumonisin diets for 100 days were similar and were not apparently altered by exposure to these mycotoxins. These results suggest that Sa/So ratios in urine, but not in hair of mink can serve as an early indicator of exposure to fumonisins in this species.     

Duckling toxicity and the production of fumonisin and moniliformin by isolates in the A and E mating populations of Gibberella fujikuroi (Fusarium moniliforme)

Two biological species of Gibberella fujikuroi (A and F mating populations) share the Fusarium moniliforme anamorph. Twenty strains of each of these biological species were tested for the ability to produce fumonisins B1, B2, and B3 and moniliformin and for toxicity to 1-day-old ducklings. Most of the members of the A mating population (19 of 20 strains) produced more than 60 micrograms of total fumonisins per g, whereas only 3 of 20 members of the F mating population produced more than trace levels of these toxins and none produced more than 40 micrograms of total fumonisins per g. In addition, only 3 of 20 members of the A mating population produced more than 1 microgram of moniliformin per g (and none produced more than 175 micrograms/g), while all 20 strains of the F mating population produced more than 85 micrograms of this toxin per g and 1 strain produced 10,345 micrograms/g. The duckling toxicity profiles of the strains of the two mating populations were similar, however, and the level of either toxin by itself was not strongly correlated with duckling toxicity. On the basis of our data we think that it is likely that the members of both of these mating populations produce additional toxins that have yet to be chemically identified. These toxins may act singly or synergistically with other compounds to induce the observed duckling toxicity.     

Trichothecene 3-O-acetyltransferase protects both the producing organism and transformed yeast from related mycotoxins. Cloning and characterization of Tri101

Trichothecene mycotoxins such as deoxynivalenol, 4,15-diacetoxyscirpenol, and T-2 toxin, are potent protein synthesis inhibitors for eukaryotic organisms. The 3-O-acetyl derivatives of these toxins were shown to reduce their in vitro activity significantly as assessed by assays using a rabbit reticulocyte translation system. The results suggested that the introduction of an O-acetyl group at the C-3 position in the biosynthetic pathway works as a resistance mechanism for Fusarium species that produce t-type trichothecenes (trichothecenes synthesized via the precursor trichotriol). A gene responsible for the 3-O-acetylation reaction, Tri101, has been successfully cloned from a Fusarium graminearum cDNA library that was designed to be expressed in Schizosaccharomyces pombe. Fission yeast transformants were selected for their ability to grow in the presence of T-2 toxin, and this strategy allowed isolation of 25 resistant clones, all of which contained a cDNA for Tri101. This is the first drug-inactivating O-acetyltransferase gene derived from antibiotic-producing organisms. The open reading frame of Tri101 codes for a polypeptide of 451 amino acid residues, which shows no similarity to any other proteins reported so far. TRI101 from recombinant Escherichia coli catalyzes O-acetylation of the trichothecene ring specifically at the C-3 position in an acetyl-CoA-dependent manner. By using the Tri101 cDNA as a probe, two least overlapping cosmid clones that cover a region of 70 kilobase pairs have been isolated from the genome of F. graminearum. Other trichothecene biosynthetic genes, Tri4, Tri5, and Tri6, were not clustered in the region covered by these cosmid clones. These new cosmid clones are considered to be located in other parts of the large biosynthetic gene cluster and might be useful for the study of trichothecene biosynthesis.     

Production of [14C]fumonisin B1 by Fusarium moniliforme MRC 826 in corn cultures

Kinetics of growth and fumonisin production by Fusarium moniliforme MRC 826 in corn "patty" cultures were investigated, and a technique was developed for the production of [14C]fumonisin B1 ([14C]FB1) by using L-[methyl-14C]methionine as the precursor. A significant (P < 0.01) correlation exists between fungal growth and FB1 (r = 0.89) and FB2 (r = 0.87) production in corn patties, beginning after 2 days and reaching the stationary phase after 14 days of incubation. [14C]FB1 was produced by adding L-[methyl-14C]methionine daily to cultures during the logarithmic phase of production. Incorporation of the isotope occurred at C-21 and C-22 of the fumonism molecule and was enhanced in the presence of unlabeled L-methionine. Although the concentration of exogenous unlabeled methionine is critical for incorporation of the 14C label, optimum incorporation was achieved by adding 50 mg of unlabeled L-methionine and 200 mu Ci of L-[methyl-14C]methionine to a corn patty (30 g) over a period of 9 days, yielding [14C]FB1 with a specific activity of 36 mu Ci/mmol.