Production of fumonisin B1 by Fusarium moniliforme NRRL 13616 in submerged culture

Fumonisin B1, a recently discovered mycotoxin, was synthesized by submerged cultures of Fusarium moniliforme NRRL 13616 grown for 29 days at 28 degrees C and 220 rpm in a basal salts medium (pH 5.0) supplemented with 90 g of glucose per liter and 3.5 g of ammonium sulfate per liter. Under these culture conditions, 74 +/- 23 micrograms of fumonisin B1 per ml was produced by 29-day-old F. moniliforme NRRL 13616 cultures. Fumonisin B1 was detected in liquid culture extracts by high-performance thin-layer chromatography. Fumonisin B1 was confirmed and quantitated by gas chromatography and gas chromatography-mass spectral analysis of the trimethylsilyl derivative. The use of a defined medium for producing fumonisin B1 in a submerged culture facilitates its isolation and provides an excellent method for conducting biosynthetic studies.     

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.     

Influence of kernel age on fumonisin B1 production in maize by Fusarium moniliforme.

Production of fumonisins by Fusarium moniliforme on naturally infected maize ears is an important food safety concern due to the toxic nature of this class of mycotoxins. Assessing the potential risk of fumonisin production in developing maize ears prior to harvest requires an understanding of the regulation of toxin biosynthesis during kernel maturation. We investigated the developmental-stage-dependent relationship between maize kernels and fumonisin B1 production by using kernels collected at the blister (R2), milk (R3), dough (R4), and dent (R5) stages following inoculation in culture at their respective field moisture contents with F. moniliforme. Highly significant differences (P 相似文献   

Effects of zinc, iron, cobalt, and manganese on Fusarium moniliforme NRRL 13616 growth and fusarin C biosynthesis in submerged cultures.

The influence of zinc, iron, cobalt, and manganese on submerged cultures of Fusarium moniliforme NRRL 13616 was assessed by measuring dry weight accumulation, fusarin C biosynthesis, and ammonia assimilation. Shake flask cultures were grown in a nitrogen-limited defined medium supplemented with various combinations of metal ions according to partial-factorial experimental designs. Zinc (26 to 3,200 ppb [26 to 3,200 ng/ml]) inhibited fusarin C biosynthesis, increased dry weight accumulation, and increased ammonia assimilation. Carbohydrate was found to be the principal component of the increased dry weight in zinc-supplemented cultures. Zinc-deficient cultures synthesized more lipid and lipidlike compounds, such as fusarin C, than did zinc-supplemented cultures. Microscopic examination showed that zinc-deficient hyphae contained numerous lipid globules which were not present in zinc-supplemented cultures. Addition of zinc (3,200 ppb) to 2- and 4-day-old cultures inhibited further fusarin C biosynthesis but did not stimulate additional dry weight accumulation. Iron (10.0 ppm) and cobalt (9.0 ppm) did not affect fusarin C biosynthesis or dry weight accumulation. Manganese (5.1 ppm) did not affect dry weight accumulation but did increase fusarin C biosynthesis in the absence of zinc. Maximum fusarin C levels, 32.3 micrograms/mg (dry weight), were produced when cultures were supplied manganese, whereas minimum fusarin C levels, 0.07 micrograms/mg (dry weight), were produced when zinc, iron, cobalt, and manganese were supplied. These results suggest a multifunctional role for zinc in affecting F. moniliforme metabolism.     

Effects of zinc, iron, cobalt, and manganese on Fusarium moniliforme NRRL 13616 growth and fusarin C biosynthesis in submerged cultures

The influence of zinc, iron, cobalt, and manganese on submerged cultures of Fusarium moniliforme NRRL 13616 was assessed by measuring dry weight accumulation, fusarin C biosynthesis, and ammonia assimilation. Shake flask cultures were grown in a nitrogen-limited defined medium supplemented with various combinations of metal ions according to partial-factorial experimental designs. Zinc (26 to 3,200 ppb [26 to 3,200 ng/ml]) inhibited fusarin C biosynthesis, increased dry weight accumulation, and increased ammonia assimilation. Carbohydrate was found to be the principal component of the increased dry weight in zinc-supplemented cultures. Zinc-deficient cultures synthesized more lipid and lipidlike compounds, such as fusarin C, than did zinc-supplemented cultures. Microscopic examination showed that zinc-deficient hyphae contained numerous lipid globules which were not present in zinc-supplemented cultures. Addition of zinc (3,200 ppb) to 2- and 4-day-old cultures inhibited further fusarin C biosynthesis but did not stimulate additional dry weight accumulation. Iron (10.0 ppm) and cobalt (9.0 ppm) did not affect fusarin C biosynthesis or dry weight accumulation. Manganese (5.1 ppm) did not affect dry weight accumulation but did increase fusarin C biosynthesis in the absence of zinc. Maximum fusarin C levels, 32.3 micrograms/mg (dry weight), were produced when cultures were supplied manganese, whereas minimum fusarin C levels, 0.07 micrograms/mg (dry weight), were produced when zinc, iron, cobalt, and manganese were supplied. These results suggest a multifunctional role for zinc in affecting F. moniliforme metabolism.     

Effects of temperature and incubation period on production of fumonisin B1 by Fusarium moniliforme.

The kinetics of the production of fumonisin B1 (FB1) by Fusarium moniliforme MRC 826 in corn cultures was investigated as a function of fungal growth at various incubation temperatures. The growth rate of F. moniliforme, as measured by ergosterol concentration, was higher at 25 degrees C than at 20 degrees C, reaching a stationary phase after 4 to 6 weeks in both cases. FB1 production commenced after 2 weeks during the active growth phase, continued to increase during the stationary phase, and decreased after 13 weeks. The overall maximal yield of FB1 (17.9 g/kg, dry weight) was obtained in corn cultures incubated at 20 degrees C for 13 weeks, but it was not significantly (P greater than 0.05) higher than the maximum yield (16.5 g/kg, dry weight) obtained at 25 degrees C after 11 weeks. However, a significantly (P less than 0.05) higher mean yield was detected at 25 degrees C (9.5 g/kg, dry weight) than at 20 degrees C (8.7 g/kg, dry weight). Production reached a plateau after 7 weeks of incubation at 25 degrees C or 9 weeks of incubation at 20 degrees C. The maximal production of FB1 at 30 degrees C was very low (0.6 g/kg, dry weight). FB1 was also found to be heat stable, as there was no reduction in the FB1 concentration after boiling culture material of F. moniliforme MRC 826.     

Effects of temperature and incubation period on production of fumonisin B1 by Fusarium moniliforme

The kinetics of the production of fumonisin B1 (FB1) by Fusarium moniliforme MRC 826 in corn cultures was investigated as a function of fungal growth at various incubation temperatures. The growth rate of F. moniliforme, as measured by ergosterol concentration, was higher at 25 degrees C than at 20 degrees C, reaching a stationary phase after 4 to 6 weeks in both cases. FB1 production commenced after 2 weeks during the active growth phase, continued to increase during the stationary phase, and decreased after 13 weeks. The overall maximal yield of FB1 (17.9 g/kg, dry weight) was obtained in corn cultures incubated at 20 degrees C for 13 weeks, but it was not significantly (P greater than 0.05) higher than the maximum yield (16.5 g/kg, dry weight) obtained at 25 degrees C after 11 weeks. However, a significantly (P less than 0.05) higher mean yield was detected at 25 degrees C (9.5 g/kg, dry weight) than at 20 degrees C (8.7 g/kg, dry weight). Production reached a plateau after 7 weeks of incubation at 25 degrees C or 9 weeks of incubation at 20 degrees C. The maximal production of FB1 at 30 degrees C was very low (0.6 g/kg, dry weight). FB1 was also found to be heat stable, as there was no reduction in the FB1 concentration after boiling culture material of F. moniliforme MRC 826.     

Developmental effects of fumonisin B1-containingFusarium moniliforme culture extract in CD1 mice

Pregnant Charles River CD1 mice were treated with a semipurified extract ofFusarium moniliforme culture containing 0, 12.5, 25, 50 or 100 mg FB₁/kg each day orally (diluted in distilled water) between gestational days (GD) 7 and 15 to evaluate the developmental toxicity of FB₁. Following sacrifice of dams on GD 18, litters were examined for gross abnormalities and divided equally for skeletal or visceral examination by routine techniques. Significant maternal mortality was observed at doses of 50 and 100 mg FB₁/kg. Dose-dependant decreases in maternal body weight gains, number of live offsprings per litter, and mean body weight of the offspring were produced at FB₁ doses of 25 mg/kg or higher. The percentage of implants resorbed increased at all doses in a dose-dependant manner. A dose-dependant increase, except at the lowest dose tested, in the incidence of ossification deficits involving digits and sternum, short and wavy ribs, and hydrocephalus of lateral and third ventricles was also evident. Cleft palate was seen only at the highest FB₁ dose. Maternal intoxication manifested as a dose-dependant increase in the severity of ascites associated mainly with increased histopathologic scores reflecting hepatocellular damage at day 18. Concommittant increases in serum alanine amino transferase (ALT) on GD 12, reflecting parenchymal liver cell damage, was also observed at all doses above 12.5 mg of FB₁/kg. These results suggest that FB₁-containingF. moniliforme culture extract is developmentally toxic in mice, and that this toxicity may be mediated by maternal hepatotoxicity.     

Effects of Fusarium moniliforme culture extracts and fumonisin B1 on DNA,RNA and protein synthesis by baby hamster kidney cells

Baby hamster kidney cells (BHK-21) were exposed to culture filtrates of 4 Fusarium moniliforme isolates containing varying levels of fumonisin B1 (FMB1) and the effects upon RNA, DNA and protein synthesis were monitored. Cells were also grown on medium amended with FMB1 only for comparison. After 24 h incubation FMB1 (100 μg/100 ml medium) reduced protein synthesis by 4% and by 18% after 48 h. Culture filtrates containing the highest levels of FMB1 also caused the greatest inhibition in protein synthesis after 24 h but after 48 h protein synthesis levels were the same as controls even though the FMB1 level was 360 μg/100 ml. Only FMB1 reduced DNA synthesis, by 8% after 24 h but after 48 h DNA levels had increased by 40 % over controls. The culture filtrates containing the highest levels of FMB1 (360 μg/100 ml) reduced DNA synthesis more than 50% after 24 h and 48 h. Culture filtrates containing lesser amounts of FMB1 in some instances stimulated DNA synthesis and inhibited it in others. There was also no correlation in the level of FMB1 with the inhibition of RNA synthesis by BHK cells. It appears that metabolites other than fumonisin produced by F. moniliforme in culture can affect and both stimulate and inhibit RNA, DNA and protein synthesis by BHK cells. This revised version was published online in June 2006 with corrections to the Cover Date.     

Production of Alanine by Fusarium moniliforme

Fusarium moniliforme grown in a chemically defined medium in submerged culture accumulated amino acids extracellularly. Alanine and glutamic acid were present in greatest amounts, with traces of glycine, lysine, threonine, and valine detectable. Increasing the glucose and urea concentrations of the medium increased yields of alanine. Further increases in alanine production occurred with elevated levels of mineral salts in the medium, whereas the addition of a vitamin mixture proved to be inhibitory. Chemical changes resulting from the growth of F. moniliforme in the final fermentation medium disclosed maximal alanine production, mycelial weight, and glucose consumption after 72 hr of incubation at 28.5 C. Total soluble nitrogen, by contrast, was minimal at the same time period. The pH remained in the alkaline range throughout the fermentation.     

Production of vomitoxin on corn by Fusarium graminearum NRRL 5883 and Fusarium roseum NRRL 6101.

Two vomitoxin-producing isolates of Fusarium spp. were grown on cracked corn for 1 to 8 weeks at 15, 20, 25, 28, and 32 degrees C. Maximum production of vomitoxin by Fusarium graminearum Schw. NRRL 5883 occurred at 30 degrees C and 40 days, and that by Fusarium roseum Schw. NRRL 6101 occurred at 26 degrees C and 41 days. These optimum production points were determined from response surface contour graphs in relation to temperature and time. Only small amounts of vomitoxin were produced at 15 and 20 degrees C by each strain. A 133-microgram quantity of vomitoxin, with an indicated purity of 95%, was isolated per gram of corn fermented with F. graminearum NRRL 5883.     

A new fumonisin from solid cultures of Fusarium moniliforme

A new fumonisin has been isolated from Fusarium moniliforme isolate MRC826 grown on corn. It was shown by NMR and mass spectrometry to be an isomer of fumonisin B₂ that has free hydroxyl groups at C-3 and C-10 instead of the normal C-3 and C-5. This new fumonisin was detected in cultures of most isolates of F. moniliforme that were examined and was usually present at concentrations similar to those of fumonisin B₂. Two isolates of F. moniliforme that produce significantly higher levels of this new isomer were identified.Abbreviations ELEM equine leukoencephalomalacia Mention of companies or products by name does not imply their endorsement by the US Department of Agriculture over others not cited.     

De novo induction of adventitious roots in excised shoots of tomatoes by fumonisin B1, a metabolite of Fusarium moniliforme

The de novo induction of roots in tomatoes (Lycopersicon esculentum) Mill. cvs. Earlypak-7, Ace, Better Boy, Roma, and Parks' Whopper by fumonisin B₁, a mycotoxin produced by Fusarium moniliforme J. Sheld., was studied. In graded dosages of fumonisin B₁, detached stems of the cultivars Ace, Better Boy, and Roma were induced to produce calluses and roots earlier than controls. The cultivar Ace was especially responsive to this mycotoxin, and following a single application, callus initiation was observed to occur within a 24–48-h period and roots were produced as early as 72 h with 10 g/shoot or as late as 96 h with low dosages. The control plants of all cultivars were completely negative for a rooting response during this time. Some cultivars treated with fumonisin B₁ showed either no response or developed signs of phytotoxicity. Those cultivars that were stimulated to produce roots did not show signs of phytotoxicity, except at dosages of 0.5 mg/plant and higher. One cultivar did not show any signs of phytotoxicity nor was it induced to root. The ability of fumonisin B₁ to affect the accumulation of calcium in other systems, and its structural similarity to sphingosine suggest that the induction of adventitious roots may be a calcium-dependent process.     

Production of fumonisin B1 and B2 byFusarium moniliforme isolated from Korean corn kerneis for feed

The natural occurrence of fumonisin B₁ (FB₁) and B₂ (FB₂), a promoter for hepatocarcinogenesis, was investigated in Korean corn kernels for feed by HPLC with fluorescence detection. From the 12 corn kernel samples, FB₁ was detected in 5 samples at levels ranging from 53 to 1327ng/g, while FB₂ was found in 4 samples from 69 to 680ng/g. In the positive samples, the average concentrations of FB₁ and FB₂ were 506 and 288ng/g, respectively. One sample (No. K3) showed the highest FB₁ and FB₂ contents as 1327 and 680ng/g, respectively. In the micrological survey on 5 positive samples for FB₁ and FB₂, 6 strains ofFusarium monlliforme were isolated, and all these isolates had a producibility of FB₁ and FB₁, with maximum levels of 80.7 to 180.9 g/g.Thisis the first report on the natural co-contamination of FB₁ and FB₂ in Korean corn kernels for feed, and on the ability ofF. moniliforme isolated from corn kernels for feed in Korea to produce FB₁ and FB₂.     

In vivo effects of fumonisin B1-producing and fumonisin B1-nonproducing Fusarium moniliforme isolates are similar: Fumonisins B2 and B3 cause hepato- and nephrotoxicity in rats

Fumonisins are mycotoxins produced by Fusarium moniliforme, F. proliferatum, and related Fusarium species found on corn. They occur naturally in corn-based feeds and foods and are suspected human esophageal carcinogens. Fumonisin B₁ (FB₁), the most common homologue, causes the animal diseases associated with F. moniliforme. Hepato- and nephrotoxicities, disrupted sphingolipid metabolism, and liver cancer have been found in rats fed FB₁. To determine the in vivo effects of diets containing fumonisins B₂ (FB₂) or B₃ (FB₃), male rats were fed culture materials (CM) of FB₁ non-producing F. moniliforme isolates to provide low (4.6–6.7 ppm), mid (32–49 ppm) or high (219–295 ppm) dietary levels of either FB₂ (FB₂CM) or FB₃ (FB₃CM). Other groups were fed culture material of an FB₁ producing isolate (FB₁CM) providing 6.9, 53 or 303~ppm total fumonisins (FB₁ : FB₂ : FB₃ = 1.0 : 0.38 : 0.15) and a tenth group was fed a control diet having no detectable fumonisins. One-half (n = 5/group) the animals were killed after three weeks, at which time the toxicological and histopathological effects of the three culture materials were similar, mimicked the effects of FB₁, and included decreased body weight gains, serum chemical indicators of hepatotoxicity, decreased kidney weights, and apoptosis of hepatocytes and kidney tubular epithelium. FB₁CM, FB₂CM, and FB₃CM affected sphingolipids, causing increased sphinganine to sphingosine ratios (Sa/So) in both liver and kidneys. The remaining animals (n = 5/group) were fed a control diet for three additional weeks. All body weight and tissue specific effects, including increased Sa/So, induced by the FB₂CM, FB₃CM and low level FB₁CM diets were absent following the recovery period. Except for mild biliary lesions found in the high dose FB₁CM group and a few apoptotic hepatocytes present in one mid- and two high-dose FB₁CM rats, no evidence of toxicity remained in these groups following the recovery period.This revised version was published online in October 2005 with corrections to the Cover Date.     

Comparative studies of hepatotoxicity and fumonisin B1 and B2 content of water and chloroform/methanol extracts of Fusarium moniliforme strain MRC 826 culture material

Fusarium moniliforme has been associated with several diseases including equine leukoencephalomalacia, human esophageal cancer and hepatotoxicity/hepatocarcinogenicity in laboratory animals. The potential health risks to animals and humans posed by F. moniliforme contaminated grains cannot be assessed until the toxins are identified and toxicologically evaluated. As part of a systematic approach to identifying the hepatotoxins produced by F. moniliforme, diets containing aqueous and chloroform/methanol (11) extracts of F. moniliforme strain MRC 826 culture material (CM) and/or the extracted CM residues were fed to male Sprague-Dawley rats for four weeks. Serum alanine aminotransferase, aspartate aminotransferase and alkaline phosphatase activities were increased after two and four weeks and microscopic liver lesions were found in those animals fed aqueous CM extract and the CM residue after chloroform/ methanol extraction. Fumonisins B₁ and B₂ were extracted from the CM by water, but not chloroform/ methanol, and were present in the toxic diets at concentrations of 93–139 and 82–147 ppm, respectively. Nontoxic diets contained 22 ppm fumonisin B₁ and 65 ppm fumonisin B₂.Abbreviations CM culture material - ELEM equine leukoencephalomalacia Mention of a trademark, proprietory name or vendor does not imply its approval by the US Department of Agriculture to the exclusion of others that may also be suitable.     

Moniliformin, a metabolite of Fusarium moniliforme NRRL 6322: purification and toxicity.

Fusarium moniliforme NRRL 6322 produced about 600 mg of recoverable moniliformin, a mycotoxic metabolite, per kg of corn grit medium. The moniliformin was extracted from the grits with methanol, purified by preparative thin-layer chromatography, and crystallized from ether. The 50% lethal dose for chicken embryos was 2.8 microgram per egg. For 1-day-old chicks dosed with moniliformin by crop intubation and for female and male mice injected intraperitoneally, the 50% lethal doses were 5.4, 20.9, and 29.1 mg per kg of body weight, respectively. The toxin did not cause a reaction on mouse skin.     

D-泛解酸内酯水解酶产生菌的筛选及产酶条件研究

筛选到一株产D-泛解酸内酯水解酶的菌株,经鉴定为串珠镶孢霉菌（Fusarium monili-forme)SW-902。产酶条件研究表明,用甘油作碳源,蛋白胨作氮源,初始pH8.0,温度26℃,摇瓶培养3d,产酶量最高,在60L发酵罐中通风发酵45h,产菌丝体生物量7.18g干菌体/L,D-泛解酸内酯水解酶酶活力达到0.92IU/g干菌体。     

D-泛解酸内酯水解酶产生菌的筛选及产酶条件研究

筛选到一株产D 泛解酸内酯水解酶的菌株 ,经鉴定为串珠镶孢霉菌 (Fusariummonili forme)SW 90 2。产酶条件研究表明 ,用甘油作碳源 ,蛋白胨作氮源 ,初始pH8 0 ,温度 2 6℃ ,摇瓶培养 3d ,产酶量最高。在 6 0L发酵罐中通风发酵 45h ,产菌丝体生物量 7 1 8g干菌体 L ,D 泛解酸内酯水解酶酶活力达到 0 92IU g干菌体