Production of fumonisins byFusarium species of Taiwan

Twenty-nineFusarium species isolated from various sources in different districts of Taiwan were tested for their ability to produce fumonisins in corn cultures. OnlyFusarium moniliforme produced fumonisin B₁ (FB₁) and fumonisin B₂ (FB₂). The finding that the other 28Fusarium species produced neither FB₁ nor FB₂ is preliminary because only one strain per species was studied. The detection of FB₁ and FB₂ in cultures ofF. moniliforme was demonstrated by TLC and HPLC, and FB₁ was further confirmed by mass spectrometry. In a separate experiment, in which 38 strains ofF. moniliforme were tested for fumonisins, approximately 66% (25/38) produced FB₁ and/or FB₂. Of the 25 strains, 14 produced only FB₁ and 11 produced both FB₁ and FB₂, and the amounts of FB₁ and FB₂ produced by different strains varied greatly. This is the first report that fumonisins are found in corn cultures experimentally infected withF. moniliforme strains from Taiwan. It is safe to assume that fumonisin producing strains ofF. moniliforme are widely distributed among the economic crops such as corn, rice, sugarcane, and sorghum throughout the Island.Abbreviations FB₁ Fumonisin B₁ - FB₂ Fumonisin B₂ - OPA o-phthalidialdehyde     

A limited survey of fumonisins in corn and corn-based products in Asian countries

Natural occurrence of fumonisins B₁ (FB₁) and B₂ (FB₂), a promoter for hepato-carcinogenesis, was investigated in corn and corn — based products sampled in Japan, Nepal, and China by high — performance liquid chromatographic method. From the 9 imported corn kernel and 6 gluten feed samples, FB₁ was detected in 8 corn (0.6 ～ 4.1μg/g) and all gluten feed (0.3 ～ 2.4μg/g) samples, while FB₂ was found in the same corn (0.3 ～ 10μg/g) and 3 gluten feed (0.8 ～ 8.5μg/g) samples. ELISA analysis also revealed the contamination of aflatoxin B₁ in 2 corn and all gluten feed samples along with fumonisins. Of 17 corn grit samples, 14 and 5 samples were contaminated with fumonisin B₁ and B₂, with maximum levels of 2.6 and 2.8μg/g, respectively. As for corn-based foodstuffs marketed in Japan, no significant contamination of fumonisins was observed. Among 24 corn kernel samples in Nepal, 12 and 7 samples were positive for FB₁ and FB₂, and averaged to 0.6 and 1.6μg/g, respectively. One sample showed the highest fumonisin contents as 4.6 and 5.5μg/g, respectively. In corn samples harvested at Shanghai and Beijing, China, FB₁ and FB₂ were detected in various concentrations. Mycological survey has also revealed the presence of a fumonisin — producing fungus in a crop field of Japan. These findings have for the first time demonstrated high levels of contamination of fumonisins in corn and corn — based products in Asian countries. Natural co — occurrence of fumonisins and aflatoxin B₁ was also detected in raw materials for mixed feed.     

Natural occurrence of fumonisins in corn puff in Argentina

Maize presents a substantial contamination by fumonisins (FB) throughout the world. In particular, Argentina presents a high frequency of positive samples and levels of up to 10,000 μg/kg of total FB, depending on the harvest campaign studied. Corn puff, an extruded product of high consumption among children and adolescent populations, was analyzed with the aim of determining the presence and levels of fumonisins B₁, B₂ and B₃. In the present study, 20 samples of different brands were collected from commercial stores. Two solvent systems, two types of agitation, and the number of extractions were tested in order to find the most suitable method to establish the level of fumonisins in corn puff. Extracts were analyzed by high performance liquid chromatography (HPLC) with fluorescence detection. A C18 column and precolumn derivatization with orthophtalaldialdehyde were utilized. The average of fumonisins found in 20 samples was 257.5 μg/kg (n?=?19), 70.4 μg/kg (n??=?14) and 73.3 μg/kg (n??=?6) for FB₁, FB₂ and FB₃, on positive samples, respectively. These are the first analyses in Argentina of this type of product, highlighting the need to continue the studies in the processing industries because, despite the extrusion process that the raw material suffers, a sample was found with a contamination of 1,649 μg/kg total fumonisins.     

Production of fumonisins B1, B2 and B3 byFusarium proliferatum Isolated from rye grains

Using the seed- plate technique, we have isolated a strain ofF. proliferatum from rye grains that produces 3 fumonisins, fumonisin B₁ (FB₁), FB₂ and FB₃ on inoculated rice and corn. Inoculated corn and rice were extracted with an aqueous methanol solution and fumonisin concentrations estimated using high performance liquid chromatography. Production of all 3 fumonisins (FB₁, FB₂ and FB₃) was much higher on rice than corn; ranging from 3816, 1068 and 985 ppm to 1643, 350 and 162 ppm respectively. We conclude that all natural substrates whereF. proliferatum is present as a component of the mycoflora should be monitored for the presence of fumonisins.     

Studies on extraction of fumonisins from rice, corn-based foods and beans

Different solvent mixtures were examined for extraction of fumonisins from various naturally contaminated and spiked foods and foodstuffs: rough rice, retail rice, rice flour, white corn flour, corn meal, corn starch, corn flakes, tortilla/corn chips, white bean flour, white beans, mung beans, adzuki beans and infant cereals. Most of the naturally contaminated samples were analyzed using the extraction solvent mixtures methanol-acetonitrile-water (25:25:50) (solvent A) and methanol-water (75:25 or 80:20) (solvents B, BB); some were extracted with 0.1 M sodium hydrogen phosphate-acetonitrile (1:1, adjusted to pH 3.0 with o-phosphoric acid) (solvent C) and methanol-0.025 M borate buffer (3:1, adjusted to pH 9.2 with 1 N sodium hydroxide) (solvent D). A 1-ml SAX solid phase extraction column was used for the cleanup in all cases except for infant cereals, for which immunoaffinity chromatography was used; fumonisin concentrations were determined by liquid chromatography. Solvent A gave slightly better extraction of fumonisins from one of two samples of naturally contaminated rough rice than solvent B (fumonisin B₁: 4080 ng/g versus 3150 ng/g; fumonisin B₂:1100 ng/ g versus 922 ng/g) and much better extraction than solvent C (1210 ng/g fumonisin B₁ and 315 ng/g fumonisin B₂) or solvent D (372 ng/ g fumonisin B1 and 191 ng/g fumonisin B2). However, spike recoveries on a similar rice naturally contaminated at a lower level were only in the 43–53% range (solvent A). Recovery of fumonisins was very poor from spiked white rice flour but satisfactory from other rice foods. Solvent A similarly gave slightly better extraction of fumonisins from a sample of naturally contaminated white corn flour than solvent B (fumonisin B₁ 1260 ng/g versus 931 ng/g; fumonisin B₂: 511 ng/g versus 447 ng/g ) and better extraction than solvents C and D. Solvent A was also a better solvent for extraction of fumonisins from naturally contaminated tortilla chips and infant cereals. Study of naturally contaminated corn starch was confounded by instability of fumonisins in this food. Recovery of fumonisins from spiked corn meal, tortilla chips, corn flakes, various types of beans and infant cereals with solvent A and/or solvent B (or BB) was satisfactory.     

Determination of fumonisins in corn: Evaluation of two purification procedures

The performance of two solid phase extraction (SPE) purification procedures, used in the determination of fumonlsin B₁ (FB₁), B₂ (FB₂) and B₃ (FB₂) In corn, was evaluated using both thin-layer chromatography (TLC) and high performance liquid chromatography (HPLC). Fewer interferences were observed In extracts prepared using the strong anion exchange (SAX) media, in contrast to those purified on C₁₈ media, where on occasions, visual discernment of the TLC bands was hampered by the presence of interfering compounds. Precipitate formation, resulting In the blocking of SPE cartridges was also encountered when using the C₁₈ procedure. HPLC analyses of extracts prepared by both media indicated that they gave comparable fumonlsin recoveries from naturally contaminated corn samples. The results suggest that the C₁₈ procedure, originally developed for the TLC analyses of FB₁ in mixed feeds, may also be applied to the determination of FB₂ and FB₂. However, where TLC is used quantitatively for fumonlsin levels <1 μg/g, purification of sample extracts on SAX media is recommended.     

Effect of<Emphasis Type="Italic">in vitro</Emphasis> digestion on fumonisin B<Subscript>1</Subscript> in corn flakes

Low levels of fumonisins have been found frequently in corn based breakfast cereals and can occur bound to protein and other matrix components.In vitro digestion of two samples of corn flakes was carried out under "fed conditions." Fumonisins were measured as o-phthaldialdehyde/mercaptoethanol derivatives by LC-fluorescence. One sample of corn flakes (FN12) had high concentrations of fumonisin B₁ (FB) (average 125 ng/g) and total bound FB₁, (TB FB₁) (average 92 ng/g) and the other (FN11) had a low level of free FB₁ (average 29 ng/g) and no detectable TB FB₁. After incubation of the samples with gastrointestinal tract solutions simulating saliva plus stomach and duodenal juices, chyme was analysed for FB₁, hydrolyzed FB₁ (HFB₁) and partially hydrolyzed fumonisin B₁ (PHFB₁). The bioaccessibility (percentage of FB₁ released from corn flakes into chyme) was 38-78% for incurred FB₁ in FN12, 8-54% for incurred plus spiked FB₁ in FN12, and 19-66% for incurred plus spiked FB₁ in FN11. HFB₁ and PHFB₁ were not detected. If free FB₁ was first extracted from sample FN12, no FB1 was detected in the chyme, indicating no contribution from TB FB₁. Concentrations were corrected for method recovery of FB₁ or, for bound FB₁, partial method recovery of HFB₁ Presented at the XIIth IUPAC International Symposium on Mycotoxins and Phycotoxins, Istanbul, Turkey, 21–25 May, 2007     

Fumonisin production by <Emphasis Type="Italic">Fusarium</Emphasis> species isolated from freshly harvested corn in Iran

Fifty-one strains of Fusarium verticillioides and F. proliferatum isolated from corn collected from four different geographic areas in Iran, namely Fars, Khuzestan, Kermanshah and Mazandaran (an endemic oesophageal cancer (OC) area) were evaluated for their ability to produce fumonisins B₁ (FB₁), B₂ (FB₂) and B₃ (FB₃) in corn culture. Fumonisin levels were determined by high-performance liquid chromatography. All tested strains of F. verticillioides and F. proliferatumproduced fumonisins within a wide range of concentrations, 197–9661 g/g, 18–1974 g/g, and 21–1725 g/g for FB₁, FB₂, and FB₃, respectively. The highest mean concentrations of FB₁, FB₂, and FB₃ were 3897, 806 and 827 g/g, respectively. Overall, 61% of the F. verticillioides and F. proliferatum strains produced higher levels of FB₃ than FB₂. The mean ratios of FB₁:FB₂, FB₁:FB₃ and FB₁:total fumonisins were 8, 7 and 0.7 for F. verticillioides and 5.7, 10.7 and 0.7 for F. proliferatum, respectively. Significant differences in some of the meteorological data (rainfall, relative humidity and minimum temperature) from the four provinces were observed. Fumonisin levels produced by F. verticillioides strains isolated from Khuzestan province (tropical zone) were significantly (P < 0.01) higher than the other three provinces. This is the first report of the fumonisin-producing ability of F.verticillioides and F. proliferatum strains isolated from corn harvested from different geographic areas in Iran.     

Fumonisins: Isolation,chemical characterization and biological effects

The fumonisin B mycotoxins (FB₁ and FB₂) have been purified and characterized from corn cultures of Fusarium moniliforme strain MRC 826. Fumonisin B₁ (FB₁, the major fumonisin produced in culture, has been shown to be responsible for the major toxicological effects of the fungus in rats, horses and pigs. Recent investigations on the purification of compounds with chromatographic characteristics similar to FB₁ have led to the identification of two new fumonisins, FB₃ and FB₄. Fumonisins A₁ and A₂, the N-acetyl derivatives of FB₁ and FB₂ respectively, were also purified and shown to be secondary metabolites of the fungus. Short-term carcinogenesis studies in a rat liver bioassay indicated that over a period of 15 to 20 days, at dietary levels of 0.05–0.1%, FB₂ and FB₃ closely mimic the toxicological and cancer initiating activity of FB₁ and thus could contribute to the toxicological effects of the fungus in animals. In contrast, no biological activity could be detected for FA₁ under identical experimental conditions. These studies and others have indicated that the fumonisin B mycotoxins, although lacking mutagenicity in the Salmonella test or genotoxicity in the DNA repair assays in primary hepatocytes, appear to induce resistant hepatocytes similar to many known hepatocarcinogens.     

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.     

Quantitation of fumonisins in corn by HPLC with o-phthalaldehyde postcolumn derivatization and their identification by LC/MS

Fumonisins B₁(FB₁) and B₂(FB₂) were isocratically separated on a fluorocarbon column without using an ion pair reagent and nonvolatile buffer during the HPLC and were detected by an o-phthalaldehyde postcolumn derivatization system using a fluorescence detector. The minimum detectable concentrations of FB₁ and FB₂ in corn by this system were 0.01 μg/g and 0.01 μg/g, respectively. The separated fumonisins were further identified by a directly interfaced ion trap MS using electrospray ionization. FB₁ and FB₂ in naturally contaminated corn were identified in the selective ion monitoring mode at concentrations of 3.75μg/g and 1.44 μg/g, respectively. This revised version was published online in August 2006 with corrections to the Cover Date.     

Mycoflora and fumonisin-producing strains ofFusarium moniliforme in mixed poultry feeds and component raw material

In a survey of the mycoflora and mycotoxins in foods and feeds, 66 samples of mixed poultry feeds and some component raw materials were investigated. Fungal counts ranged from < 10² to 1.3 × 10⁶ CFU/g.Fusarium spp. counts ranged from 10² to 1.0 × 10⁶ CFU/g. TheFusarium spp. strains isolated were screened for their potential to produce fumonisin B₁ (FB₁) and fumonisin B₂ (FB₂) in maize cultures. Samples and maize cultures were analysed for FB₁ and FB₂ using TLC and fluorescamine-derivative HPLC. No fumonisins were detected in the samples (<6 ppm).Fusarium moniliforme was isolated in 59.1% of samples, and 97.4% of the strains produced FB₁ and 79.4% of strains produced FB₂ in maize cultures. Some isolates produced higher FB₁ and FB₂ levels than the reference strainF. moniliforme MRC 826.     

Natural occurrence of Fusarium species and fumonisin-production by toxigenic strains isolated from poultry feeds in Argentina

Fusarium species and fumonisin production by toxigenic strains were investigated. During 1996–1998, 158 samples of poultry feeds were collected from a factory located in the department of Río Cuarto Córdoba province, Argentina. The most common species of Fusarium were F. moniliforme (60.7%) and F. nygamai (35.4%) followed by F. semitectum, F. subglutinans, F. proliferatum, F. dlamini, F. solani, F. oxysporum and F. napiforme. Fungal counts ranged from 1 × 10³ to 8 × 10⁵ CFU/g with mean values from 1.5 × 10³ to 2.3 × 10⁵ CFU/g. The highest counts were for F. dlamini, F. subglutinans, F. moniliforme and F. nygamai. Strains of F. moniliforme, F. nygamai, and F. proliferatum were screened for their potential to produce fumonisin B₁ (FB₁), fumonisin B₂ (FB₂) and fumonisin B₃ (FB₃) in corn grain. The samples were analysed using a modified high performance liquid chromatography method. The strains assayed, 43 strains, produced three fumonisins. There was a high degree of variability in the quantities of FB₁, FB₂, and FB₃ produced. The toxin produced in highest levels by the majority of the strains was FB₁. The range of concentration varied from 5.4 to 3,991, 1.01 to 189 and 0.4 to 765 ppm per gram of corn for FB₁, FB₂ and FB₃ respectively. The toxigenic pattern of strains was normal, although two strains of F. moniliforme produced exceptionally high concentrations of FB₃ and minor concentrations of FB₂ and FB₁. This is the first report from Argentina on Fusarium species in poultry feeds and fumonisin production by these strains.This revised version was published online in October 2005 with corrections to the Cover Date.     

Mycotoxin Production by Fusarium proliferatum isolates from rice with Fusarium sheath rot disease

Twenty samples of unpolished (rough) rice collected in Arkansas and Texas during the 1995 harvesting season from fields exhibiting Fusarium sheath rot disease or panicle blight were previously shown to include 8 samples positive for fumonisin B₁(FB₁) in the range 2.2–5.2 ppm, and moniliformin (MON), but no beauvericin (BEA), deoxynivalenol, its derivatives or zearalenone were detected. Fifteen cultures of F. proliferatum were established from the 20 rough rice samples. Single spore isolates of each culture were grown on rice and tested for the production of fumonisins (FB₁, FB₂, FB₃, etc.), MON and BEA. All 15 isolates produced FB₁, FB₂, MON and BEA in culture on rice. No deoxynivalenol, its derivatives orzearalenone were detected. Seven cultures produced FB₁ at >50ppm (range 80–230 ppm), with therest producing FB₁ in the range 14–43 ppm.FB₂ was produced in the range 5–47 ppm, and those cultures which produced the most FB₁ also produced the most FB₂. Of the 15 cultures producing MON, 11 produced it at >100 ppm in the range 188–6018 ppm, with the rest producing in the range 7–64 ppm. BEA was produced in the range 109–1350 ppm. Other derivatives of fumonisins, including FA₁, FA₂ and partially hydrolyzed FB₁, as well asseveral unknown metabolites including a compound with MW 414, were identified in culture extracts by continuous flow fast atom bombardment with ion spraymass spectrometry (CF/FAB/MS). Further study is needed to identify the factors that control production of FB₁, MON and BEA by F.proliferatu in culture and in field samples. This revised version was published online in June 2006 with corrections to the Cover Date.     

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.     

Factors affecting the growth of Fusarium proliferatum and the production of fumonisin B1: oxygen and pH

Fumonisins are mycotoxins produced primarily by Fusarium moniliforme and Fusarium proliferatum in corn. In liquid culture, production of fumonisin B₁ (FB₁), the most common moiety of the family of fumonisins, can be obtained using a defined medium that is nitrogen-limited. Under nitrogen-limited conditions both growth and the production of FB₁ were greatly influenced by pH and aeration. At pH above 5.0, F. proliferatum grew normally but produced little FB₁ (<100 μg m⁻¹). At pH below 5.0, there was less growth but substantially more FB₁. Below a pH of 2.5, both growth and metabolism were slower with very little FB₁ produced. When the optimal pH range of between 3.0 and 4.0 under well-aerated conditions was used, both growth and FB₁ production were high. However, under oxygen-limited conditions, less growth occurred, glucose consumption was increased, and no FB₁ was produced. Received 16 May 1997/ Accepted in revised form 03 September 1997     

Determination of mycobiota and mycotoxins in pig feed in central Argentina

Aims: To evaluate the mycobiota and natural levels of aflatoxins, fumonisins and zearalenone present in compound feed and home‐corn grains intended for fattening pigs. Methods and Results: Total fungi, Fusarium and Aspergillus species occurrence were examined. Aflatoxins and zearalenone were detected by thin‐layer chromatography and fumonisins by high‐pressure liquid chromatography. Fungal counts were generally higher than 1 × 10⁵ colony forming units (CFU) ml^?1. Aspergillus flavus, Aspergillus parasiticus and Fusarium verticillioides were the most prevalent species. FB₁ and FB₂ were detected in all feed and corn samples. Aflatoxin B₁ was detected in 33·33% of initial and growing feed and in 44·44% of final feed samples. It was not detected in corn samples. All feed and corn samples were negative for AFB₂, AFG₁, AFG₂ and ZEA presence during all growing stages tested. Conclusions: Fungal counts at all growing periods exceeded the levels proposed as feed hygienic quality limits. Aflatoxin levels in all feeds and fumonisin levels in many samples were higher than the established regulations. Significance and Impact of the Study: The presence of mycotoxins indicates the existence of contamination. This fact requires periodic monitoring to prevent the occurrence of mycotoxicosis in animal production, to reduce the economic losses and to minimize hazards to human health.     

Occurrence of fumonisins in maize imported into Iran during 2001–2002

Fumonisins, fungal toxins found primarily in maize and produced by various Fusarium species, have been shown to cause a variety of significant adverse health effects in livestock and experimental animals, and are probable human carcinogens. Thirty-three maize samples were collected at ports from bulk shipments, which were imported into Iran from six countries during 2001–2002, and analysed by HPLC for the most abundant of the naturally occurring fumonisin analogues, namely fumonisins B₁ (FB₁), B₂ (FB₂) and B₃ (FB₃). Of the 33 samples, 21 (64%) were found to contain FB₁ (58–512 μg/kg) at levels above 10 μg/kg. The frequency of FB₁ found in maize samples imported from Uruguay and Canada was 75%, followed by China and Argentina (67%), USA (60%), and Brazil (50%). The average FB₁ level was 266 and 169 μg/kg for positive and all samples, respectively. Medians were 250 and 146 μg/kg for positive and all samples, respectively. FB₂ levels ranged from not detected (<10 μg/kg) to 53 μg/kg, whereas no sample had an FB₃ level above the detection level (10 μg/kg). This is the first report of fumonisin contamination of imported maize in Iran. Although, the level of all detected fumonisins were below the Iranian and FDA tolerance levels for foods and feeds, It is necessary to maintain the strict rules to ensure continued safety of imported maize.     

Evalution of liquid media for fumonisin production byFusarium moniliforme MCR 826

Production of fumonlsins B₁ (FB₁) and B₂ (FB₂) by 5 lyophillzation batches ofFusarium moniliforme strain MRC 826 was studied in several liquid media and vermlculite supplemented with liquid media. In addition the effect of different parameters including pH, Inert material, shake versus stationary cultures as well as different carbon sources on the production of the fumonlsins were investigated. Fumonlsin production in liquid cultures was significantly (P<0.01) correlated (r=0.92–0.98) with fungal growth, which in turn is affected by the pH of the medium as well as the carbon source utilized. The highest FB₁ yields (approximately 40 mg/l) over the incubation period of 14 days were produced in a chemically defined medium with glucose as carbon source set at an initial pH value of 4. FB₁ production in “corn patty” cultures (approximately 1 to 3 g/kg), however, by far exceeded that obtained in the liquid media, while poor fungal growth and fumonlsin production was obtained in vermlculite supplemented cultures. From these studies it became clear that the ability of a culture to produce fumonlsins is determined by the interaction of a variety of physiological and nutritional factors regarding the inoculum and the culture medium.     

Fum3p, a 2-Ketoglutarate-Dependent Dioxygenase Required for C-5 Hydroxylation of Fumonisins in Fusarium verticillioides

Fumonisins are polyketide-derived mycotoxins produced by several agriculturally important Fusarium species. The B series fumonisins, FB₁, FB₂, FB₃, and FB₄, are fumonisins produced by wild-type Fusarium verticillioides strains, differing in the number and location of hydroxyl groups attached to the carbon backbone. We characterized the protein encoded by FUM3, a gene in the fumonisin biosynthetic gene cluster. The 33-kDa FUM3 protein (Fum3p) was heterologously expressed and purified from Saccharomyces cerevisiae. Yeast cells expressing the Fum3p converted FB₃ to FB₁, indicating that Fum3p catalyzes the C-5 hydroxylation of fumonisins. This result was verified by assaying the activity of Fum3p purified from yeast cells. The C-5 hydroxylase activity of purified Fum3p required 2-ketoglutarate, Fe²⁺, ascorbic acid, and catalase, all of which are required for 2-ketoglutarate-dependent dioxygenases. The protein also contains two His motifs that are highly conserved in this family of dioxygenases. Thus, Fum3p is a 2-ketoglutarate-dependent dioxygenase required for the addition of the C-5 hydroxyl group of fumonisins.