Incidence of Fusarium verticillioides and levels of fumonisins in corn from main production areas in Iran

A total of 52 corn samples collected in 2000 from four main corn production provinces of Iran (Fars, Kermanshah, Khuzestan, and Mazandaran) were analyzed for contamination with Fusarium verticillioides and fumonisins (FB(1), FB(2), FB(3), and 3-epi-FB(3)). The mean incidence of F. verticillioides (percent of kernels infected) for these four areas was 26.7, 21.4, 24.9, and 59.0%, respectively. The incidence in Mazandaran was significantly (p < 0.05) above that of the other areas. All samples from Mazandaran were contaminated with fumonisins with a mean level of total fumonisins of 10674 microg/kg. In contrast, the incidence of fumonisin contamination above 10 microg/kg was 53 (8/15), 42 (5/12), and 57% (8/14) in the samples from Fars, Kermanshah, and Khuzestan, respectively, and the corresponding mean total fumonisin levels were 215, 71, and 174 microg/kg, respectively. No statistical differences (p > 0.05) were observed in the fumonisin levels of the corn samples from these three provinces, which were significantly (p < 0.05) lower than the fumonisin contamination in samples from Mazandaran.     

Evaluation of conventional and organic italian foodstuffs for deoxynivalenol and fumonisins B(1) and B(2)

Two lots of human foodstuffs from conventional and organic brand foods were purchased from supermarkets and analyzed for three Fusarium toxins, deoxynivalenol, by GC-ECD, and fumonisins B(1) and B(2) (FB(1)-FB(2)), by LC-MS. The occurrence of deoxynivalenol contamination was higher than 80% in both organic and conventional foods; fumonisin B(1) was found in 20% of organic foods and in 31% of conventional ones and fumonisin B(2) in more than the 32% of the food samples from both the agricultural practices. The highest median concentration of deoxynivalenol occurred in conventional rice-based foodstuffs (207 microg/kg): that of fumonisin B(1) in conventional maize-based foods (345 microg/kg) and that of fumonisin B(2) in organic wheat-based foods (210 microg/kg).     

Mycoflora and fumonisin contamination in Brazilian corn from sowing to harvest

The present study aimed to analyze the mycoflora and potential mycotoxin contamination of soil and corn samples collected at different plant maturity stages in Cap?o Bonito and Ribeir?o Preto, two regions of the State of S?o Paulo, Brazil. In addition, the data obtained were correlated with the occurrence of wind-dispersed fungi and the predominant climatic conditions of the two regions studied. Corn mycoflora profiles showed that Fusarium verticillioides prevailed in 35% of the samples from Cap?o Bonito and in 49% of the samples from Ribeir?o Preto. Examination of wind-dispersed fungi also revealed a high incidence of F. verticillioides. Soil mycoflora analyses showed that Penicilliumwas the most prevalent genus, although F. verticillioides was present in 55.5% of Cap?o Bonito's samples and in 26.7% of Ribeir?o Preto's samples. With respect to water activity, the corn kernels most contaminated with F. verticillioides had water activity levels of 0.70-0.80. HPLC analysis of fumonisins revealed that 88.5% of Cap?o Bonito's kernels were contaminated with fumonisin B(1) (FB(1)) (0.09-10.87 microg/g) and 53.8% with fumonisin B(2) (FB(2)) (0.05-0.52 microg/g); Ribeir?o Preto's kernels presented contamination levels of 93.5% for FB(1) (0.11-17.69 microg/g) and 61.3% for FB(2) (0.05-5.24 microg/g). No aflatoxins were detected by thin-layer chromatography in corn grains of either region. The concomitant occurrence of F. verticillioides and fumonisins in most of the field corn assayed demonstrates the importance of an effective control of cultivation throughout the plant maturity stages.     

Occurrence of Fusarium spp. and fumonisin in durum wheat grains

A survey was carried out to determine Fusarium species and fumonisin contamination in 55 durum wheat (Triticum turgidum L. var. durum) samples collected during two harvest seasons (2007 and 2008) using HPLC and further LC-MS/MS confirmation. All samples showed Fusarium contamination with infection levels ranging from 8 to 66%, F. proliferatum being the species most frequently isolated during 2007 and the second most frequently isolated one during the 2008 harvest season, respectively. Natural contamination with fumonisins was found in both harvest seasons. In 2007, 97% of the samples showed total fumonisin (FB(1) + FB(2)) levels ranging from 10.5 to 1245.7 ng/g, while very low levels of fumonisins were detected in samples collected during 2008. These results could be explained by differences in the amount of rainfall during both periods evaluated. A selected number (n = 48) of F. proliferatum isolates showed fumonisin production capability on autoclaved rice. This is the first report of the presence of natural fumonisins in durum wheat grains.     

Analysis of fumonisin B(1) in Fusarium proliferatum-infected asparagus spears and garlic bulbs from Germany by liquid chromatography-electrospray ionization mass spectrometry

Fusarium proliferatum is one of a group of fungal species that produce fumonisins and is considered to be a pathogen of many economically important plants. The occurrence of fumonisin B(1) (FB(1)) in F. proliferatum-infected asparagus spears from Germany was investigated using a liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS) method with isotopically labeled fumonisin FB(1)-d(6) as internal standard. FB(1) was detected in 9 of the 10 samples in amounts ranging from 36.4 to 4513.7 ng/g (based on dry weight). Furthermore, the capability of producing FB(1) by the fungus in garlic bulbs was investigated. Therefore, garlic was cultured in F. proliferatum-contaminated soil, and the bulbs were screened for infection with F. proliferatum and for the occurrence of fumonisins by LC-MS. F. proliferatum was detectable in the garlic tissue, and all samples contained FB(1) (26.0-94.6 ng/g). This is the first report of the natural occurrence of FB(1) in German asparagus spears, and these findings suggest a potential for natural contamination of garlic bulbs with fumonisins.     

Surveillance of fumonisins in maize-based feeds and cereals from spain.

A survey has been carried out to determine the levels of fumonisins in 171 samples of maize-based feeds and cereals available in Spain. Also, the samples were examined for mold count and fungal species. Aspergillus, Penicillium, and Fusarium were the most frequent genera, and Fusarium and Aphanocladium had the highest individual percentage counts. Regarding Fusarium species, F. moniliforme (47. 4%) was the predominant species; F. proliferatum (5.3%) and F. subglutinans (7.0%) were isolated at low frequency. The high-performance liquid chromatography-o-phthaldialdehyde fluorescence method was used for the analysis of fumonisins. The highest levels of fumonisins were detected in maize. Overall, fumonisin B(1) (FB(1)) and fumonisin B(2) (FB(2)) were detected in 79.5 and 14.6%, of samples respectively, with average FB(1) levels of 3.3 microg/g and average FB(2) levels of 1.7 microg/g. Low levels of fumonisins in wheat, barley, and soybean were detected. This would appear to be the first report of concentrations of fumonisins in these commodities.     

Evaluation of fumonisin contamination in cornflakes on the Belgian market by "flow-through" assay screening and LC-MS/MS analyses

A total of 205 cornflake samples collected in Belgian retail stores during 2003-2004 were surveyed for the natural occurrence of fumonisin B1 (FB1), B2 (FB2), and B3 (FB3). These cornflake samples, originating from conventional as well as from organic production, were analyzed using an intralaboratory-validated LC-MS/MS method. Additionally, 90 cornflake samples were subjected to rapid screening using a flow-through enzyme immunoassay method to demonstrate the practicability of a screening test coupled to a validated confirmatory LC-MS/MS method for the management of food safety risks. FB(1) concentrations ranged from not detected (nd) [LOD (FB1) = 20 microg/kg] to 464 microg/kg with mean and median concentrations of respectively 104 +/- 113 and 54 microg/kg. For FB2 and FB3, the concentration ranges varied respectively from nd [LOD (FB2) = 7.5 microg/kg] to 43 microg/kg and from nd [LOD (FB3) = 12.5 microg/kg] to 90 microg/kg. Mean concentrations for FB2 and FB3 were respectively 12 +/- 8 and 21 +/- 15 microg/kg, while the median concentration was 11 microg/kg for FB2 and 19 microg/kg for FB3. From the statistical tests (chi2 and ANOVA model III), it could be concluded that the agricultural practice did not have any significant effect on the fumonisin concentrations but that the variation between different batches was significant (p < 0.0001).     

Determination of N-(carboxymethyl)fumonisin B(1) in corn products by liquid chromatography/electrospray ionization--mass spectrometry

It is well-known that fumonisin B(1) (FB(1)) in corn meal decreases during baking, frying, and cooking, but it is still not exactly clear how heating affects the formation of N-(carboxymethyl)fumonisin B(1) (NCM-FB(1)), the reaction product of FB(1) and reducing sugars. In model experiments corn grits were spiked with FB(1) (2 mg/kg) and D-glucose (50 g/kg) or sucrose (50 g/kg) and manufactured into extrusion products at various temperatures (160--180 degrees C) and moisture levels (16--20%). A liquid chromatography/electrospray ionization-mass spectrometry method using isotopically labeled fumonisin FB(1)-d(6) as an internal standard was developed for the determination of NCM-FB(1). For sample cleanup solid-phase C18 cartridges were used. The detection limit achieved with this method was 10 ng/g (signal-noise ratio = 3:1) using the protonated molecule [M + H](+) signal of NCM-FB(1) (m/z 780) in the selected ion monitoring mode. Low concentrations of NCM-FB(1) (29-97 ng/g) were detected in all samples spiked with D-glucose and FB(1), whereas those spiked with FB(1) and sucrose showed only NCM-FB(1) in samples produced at 180 degrees C (NCM-FB(1) = 27 ng/g). Various corn-containing food samples from the German market were analyzed for the presence of NCM-FB(1), FB(1), and hydrolyzed fumonisin B(1) (HFB(1)). All samples were contaminated with FB(1) (22--194 ng/g) and HFB(1) (5--247 ng/g). Six of nine samples contained NCM-FB(1) in low concentrations ranging from 10 to 76 ng/g. From these data and the low toxicity of NCM-FB(1) it can be concluded that the significance of NCM-FB(1) in food seems to be a minor one.     

Comparison of extraction buffers for the detection of fumonisin B(1) in corn by immunoassay and high-performance liquid chromatography

The Associatian of Official Analytical Chemists approved method for quantification of fumonisin B(1) (FB(1)) in corn meal or corn-based food products includes extraction into methanol (MeOH)/water (3:1, v/v). Disposal of the extraction medium can pose safety and environmental problems. To secure a rapid and inexpensive screen for FB(1) contamination, a sensitive competitive ELISA using a rabbit polyclonal antibody was developed. This assay was used in a comparative study measuring the extraction efficiency of FB(1) in aqueous or organic solvent buffers using 16 field corn samples. An aqueous phosphate buffer was found to be suitable for extracting FB(1), thus eliminating the need for organic solvents. HPLC and ELISA determinations compared well in fortified samples at known concentrations between 1 and 50 microg/mL of extract. Overestimation at levels >50 microg/mL were common. The characteristics and application of the ELISA for screening purposes are discussed.     

Mycoflora and fumonisin mycotoxins associated with cowpea [Vigna unguiculata (L.) Walp] seeds

Cowpea seed samples from South Africa and Benin were analyzed for seed mycoflora. Fusariumspecies detected were F. equiseti, F. chlamydosporum, F. graminearum, F. proliferatum, F. sambucinum, F. semitectum, and F. subglutinans. Cowpea seed from South Africa and Benin and F. proliferatum isolates from Benin, inoculated onto maize patty medium, were analyzed for fumonisin production. Samples were extracted with methanol/water and cleaned up on strong anion exchange solid phase extraction cartridges. HPLC with precolumn derivatization using o-phthaldialdehyde was used for the detection and quantification of fumonisins. Cowpea cultivars from South Africa showed the presence of fumonisin B(1) at concentrations ranging between 0.12 and 0.61 microg/g, whereas those from Benin showed no fumonisins. This is believed to be the first report of the natural occurrence of FB(1) on cowpea seed. Fumonisin B(1), B(2), and B(3) were produced by all F. proliferatum isolates. Total fumonisin concentrations were between 0.8 and 25.30 microg/g, and the highest level of FB(1) detected was 16.86 microg/g.     

Mycotoxins in corn-based food products consumed in Brazil: an exposure assessment for fumonisins

Samples from 10 different corn-based food products commercially sold in the Federal District of Brazil were analyzed for fumonisins (FB1 and FB2) using HPLC/fluorescence following naphthalene-2,3 dicarboxaldehyde (NDA) derivatization (limit of quantification (LOQ) = 0.020 mg/kg). Samples were also analyzed for aflatoxins (B1, B2, G1, and G2) on a thin-layer chromatrography (TLC) plate under UV light (LOQ of 2 microg/kg). From the 208 samples analyzed, 80.7 and 71.6% had quantifiable levels of FB1 and FB2, respectively. Mean levels of total fumonisins (FB1 + FB2) ranged from 0.127 mg/kg for corn flakes to 2.04 mg/kg for cornmeal ( creme de milho). No FBs were detected in any of the fresh, sweet corn on the cob samples analyzed. Aflatoxins were not detected in any of the 101 samples analyzed. The daily intakes of fumonisins through the consumption of corn-based food products was estimated using consumption data estimated from the 2002/2003 Brazilian Household Budget Survey and the level of fumonisins found in this and other studies conducted in Brazil. In the Federal District, the calculated total daily intake for the total and the consumers-only populations represented, respectively, 9.0 and 159% of the provisional maximum total daily intake (PMTDI) of 2 microg/kg body weight per day. At the national level, the intakes were calculated based on the fumonisin levels found in the Federal District and on published data from studies conducted elsewhere in the country. They represented 24.1 and 355% PMTDI for the total and the consumers-only populations, respectively. The high incidence of fumonisins in some corn-based products and the exposure levels found for specific subpopulations in the present study indicate the need for setting safe regulatory levels for fumonisins in food in Brazil.     

Simultaneous determination of fumonisin B(1) and hydrolyzed fumonisin B(1) in corn products by liquid chromatography/electrospray ionization mass spectrometry

A method for the simultaneous determination of fumonisin B(1) (FB(1)) and its major hydrolysis product (HFB(1)), which is known to be formed during alkaline treatment of fumonisin-containing corn meal, was devised to analyze the levels of these mycotoxins in corn products available on the German market. Liquid chromatography/electrospray mass spectrometry in combination with selected ion monitoring (SIM) was used for unambiguous detection of FB(1) and HFB(1) after extraction of samples with acetonitrile/methanol/water (25:25:50) and solid-phase C18 cleanup. Quantitation was carried out using labeled fumonisin FB(1)-D(6) as an internal standard. The detection limits achieved with this method were 8 ng/g for HFB(1) (signal-noise ratio = 5:1) and 5 ng/g for FB(1) (s/n = 5:1) using the protonated molecule signals m/z 406 and 722 in the SIM mode. A screening of several corn-containing foodstuffs, among them extrusion products and alkali-processed corn food such as tortilla chips, showed HFB(1) and FB(1) contamination with levels of 8-80 and 5-450 ng/g, respectively.     

Mold occurrence and aflatoxin B(1) and fumonisin B(1) determination in corn samples in Venezuela

Fumonisins are mycotoxins produced mainly by Fusarium moniliforme and Fusarium proliferatum, which have been associated with several animal and human diseases. Aflatoxins are hepatotoxic, mutagenic, and teratogenic metabolites produced by Aspergillus flavus and Aspergillus parasiticus. Both have been reported at high levels in corn. This study was pursued to determine mold, aflatoxin B(1) (AFTB(1)), and fumonisin B(1) (FB(1)) levels in white and yellow corn. Mold levels were determined using potato dextrose agar and identification of the main genus of molds present in corn, AFTB(1) levels by immunoaffinity chromatography, and FB(1) levels by a Bond-Elut SAX cartridge and HPLC. AFTB(1) an     

Mycotoxin contamination of dietary and medicinal wild plants in the Eastern Cape Province of South Africa

Nineteen dietary and 30 medicinal wild plants used by residents of the Eastern Cape Province of South Africa were investigated for the presence of fumonisin B1 and aflatoxin B1. The plants were extracted in water, and cleanup was undertaken on immunoaffinity cartridges; analysis was by HPLC using fluorescence detection. None of the plant extracts contained detectable levels of aflatoxin B1; however, eight plants, four dietary and four medicinal, were positive for fumonisin B1 at levels ranging from 34 to 524 microg/kg and from 8 to 1553 microg/kg, respectively. The presence of fumonisin B1 was confirmed by LC-MS/MS using positive ion electrospray ionization. Fumonisin B1 provided characteristic fragment ions at m/z 704, 686, 546, 528, 370, and 352 corresponding to sequential loss of H2O and tricarboxylic acid moieties from the alkyl backbone. These results indicate that exposure to fumonisin B1 is much more widespread than initially thought and is the first report of mycotoxin contamination in South African medicinal and dietary wild plants.     

Natural occurrence of fumonisins B(1) and B(2) in domestic maize of Taiwan.

Samples of maize grown in various districts of Taiwan were collected and analyzed for the presence of fumonisin B(1) (FB(1)) and fumonisin B(2) (FB(2)) using high-performance liquid chromatography. Forty-nine (44.5%) and 2 (1.8%) of 110 samples were found to contain FB(1) (109-1148 ng/g) and FB(2) (222-255 ng/g), respectively. The frequency of detection and also the maximum FB(1) concentration were found in samples from Penton (2/2, 262 ng/g), followed by Chiayi (18/26, 264 ng/g), Tainan (8/16, 160 ng/g), Hualinen (5/14, 1148 ng/g), Taitung (7/20, 109 ng/g), and Yunlin (9/26, 361 ng/g). Of the 110 samples examined, only 2 samples from Hualinen had been detected containing FB(2). During an analysis of the distribution pattern of FB(1), it became apparent that >79% of tested samples had FB(1) concentrations <100 ng/g, whereas 2.7% (or 3 samples) contained FB(1) >300 ng/g. These results clearly illustrated that domestically produced maize for human consumption is frequently contaminated with FB(1).     

Development of a sensitive ELISA for the determination of fumonisin B(1) in cereals

Monoclonal fumonisin B(1) antibodies with high titer were raised by using FB(1)-glutaraldehyde-keyhole limpet hemocyanin immunogen prepared by a short cross-linker reagent (glutaraldehyde). Mean cross-reactivities of the selected monoclonal antibody for FB(1), FB(2), and FB(3) were 100, 91.8, and 209%, respectively; no reactivity was found with hydrolyzed fumonisin. A direct, competitive enzyme-linked immunosorbent assay for the quantitative determination of FB(1) in cereals has been developed with this antibody. Fifty percent acetonitrile-based solvent with some additives was used for extraction of cereals, and the diluted extracts were used without cleanup in the test. The mean within-assay and interassay coefficients of variation for the standard curve were <10%. The measuring range of this test is 10-500 ng/g, with a detection limit of 7.6 ng/g FB(1). The toxin recovery from cereals infected with 50-200 ng/g of FB(1) varied between 61 and 84%. According to the comparable results of naturally infected maize samples, this test proved to be suitable for the rapid screening of food and feed samples for the presence of FBs.     

Mycoflora and occurrence of aflatoxin B(1) and fumonisin B(1) during storage of Brazilian sorghum

The present study is a 1-year follow up of the mycoflora of 140 samples of Brazilian freshly harvested (10) and stored (130) sorghum, the levels of aflatoxin and fumonisin contamination detected in the grains, and the prevailing abiotic factors (grain moisture content, water activity, temperature, relative humidity, and mean rainfall) at the time of sampling. The results show a predominance of the genera Phoma (57.1%), Aspergillus (42.7%), Fusarium (25.0%), and Rhizopus (21.4%) and the presence of nine other filamentous fungi. Fusarium, Aspergillus, and Penicillium, the three most important genera in terms of toxicity, presented numbers of colony forming units per gram of sorghum (CFU/g) that varied from 1 x 10(3) to 36 x 10(3), from 1 x 10(3) to 295 x 10(3), and from 1 x 10(3) to 20 x 10(3) CFU/g, respectively. The species most frequently found were Aspergillus flavus and Fusarium moniliforme. Of the total samples analyzed, 12.8% were contaminated with aflatoxin B(1) (concentration mean = 7-33 microg/kg) and 74.2% with fumonisin B(1) (concentration mean = 0.11-0.15 microg/g). This paper is the first report of the natural occurrence of aflatoxins and fumonisins in sorghum grain from Brazil.     

Fluorescence polarization as a means for determination of fumonisins in maize

Fumonisins, mycotoxins produced by certain species of Fusaria, are commonly found worldwide as contaminants in maize. This paper reports the development of a rapid, portable fluorescence polarization-based assay for fumonisins in maize. The assay was based on the competition of unlabeled fumonisin, from a sample, with a fluorescently tagged fumonisin (FB(1)-FL) for a fumonisin-specific monoclonal antibody in solution. The fluorescence polarization (FP) of the tagged fumonisin was increased upon binding with the antibody. In the presence of free toxin, less of the FB(1)-FL was bound and the polarization signal was decreased. The assays were very simple to perform, requiring only mixing of an aqueous extract of maize with the tagged fumonisin and antibody, and required <2 min per sample, excluding extraction time. Two permutations of the assay were tested, one with each sample matrix serving as its own blank, and the other with all of the samples compared relative to a PBS blank with normalization of the data similar to an ELISA. The limit of detection, defined as the toxin content associated with a fluorescence polarization signal 5 standard deviations from that of a fumonisin-free control, was 0.5 microg of FB(1)/g in spiked maize. Recoveries from spiked maize over the range of 0.5-20 ppm averaged 94.3 +/- 13.8%. Forty-eight samples of field-contaminated maize were tested by the FP and an established HPLC method, with a good correlation between the two (r(2) = 0.85-0.88). For these samples, the two variations of the FP assay also compared well to one another (r(2) = 0.97), suggesting the assay principle is very robust. The results, combined with the speed and ease of use for the assay, suggest that this technology has substantial potential as a screening tool for mycotoxins in foods.     

Effect of essential oils on the growth of Fusarium verticillioides and fumonisin contamination in corn

Essential oils extracted by hydrodistillation from local plants in Benin, western Africa, and oil from seeds of the neem tree (Azadirachta indica) were evaluated in vitro and in vivo for their efficacy against Fusarium verticillioides infection and fumonisin contamination. Fumonisin in corn was quantified using a fluorometer and the Vicam method. Oils from Cymbopogon citratus, Ocimum basilicum, and Ocimum gratissimum were the most effective in vitro, completely inhibiting the growth of F. verticillioides at lower concentrations over 21 days of incubation. These oils reduced the incidence of F. verticillioides in corn and totally inhibited fungal growth at concentrations of 8, 6.4, and 4.8 microL/g, respectively, over 21 days. At the concentration of 4.8 microL/g, these oils did not affect significantly fumonisin production. However, a marked reduction of fumonisin level was observed in corn stored in closed conditions. The oils adversely affected kernel germination at 4.8 microL/g and therefore cannot be recommended for controlling F. verticillioides on stored corn used as seeds, when used at this concentration. The oil of neem seeds showed no inhibitory effect but rather accelerated the growth of F. verticillioides.     

Fumonisin B-glucose reaction products are less toxic when fed to swine

The effects of fumonisin B-glucose reaction products in swine diets was examined. Pigs were fed diets containing 528 micromol of total fumonisin B/kg (FB), 528 micromol of total FB-glucose adducts/kg (FB-G, 122 micromol of unreacted FB/kg), or 0 micromol of total FB/kg for 15 days to test the efficacy of the FB-G reaction products in detoxifying FB. Weight gain in FB pigs was lower than in FB-G or controls, which was correlated with feed intake reduction in FB pigs. Serum aspartate aminotransferase, gamma-glutamyltransferase, and total bilirubin in FB pigs were higher than in FB-G or control pigs. Serum sphinganine/shingosine ratios in FB pigs were higher than in FB-G or control pigs. Microscopic examination of tissues from FB pigs showed generalized liver necrosis and apoptosis with marked cellular pleomorphism and disorganized hepatic cords. The liver and kidneys in the FB-G group appeared to be normal. Tissues of controls were free of lesions. Results suggest that dietary FB-G products are less toxic to swine and may provide an detoxification approach in instances of widespread FB grain contamination (p < 0.05).