Progress 10/01/00 to 09/30/05
Outputs
In limited previous studies of the Ascomycete fungus Gibberella zeae in North America, the populations examined were genetically and phenotypically diverse and could be viewed as subsamples of a larger population. Our objective in this study was to test the hypothesis that a homogeneous, randomly mating population of G. zeae is contiguous throughout the central and eastern United States across a span of several years. We analysed presence/absence alleles based on amplified fragment length polymorphisms (AFLPs) at 30 loci, 24 of which are defined genetically on a linkage map of G. zeae, from > 500 isolates in eight field populations from seven states collected during the 1998, 1999 and 2000 cropping seasons. All these strains had AFLP profiles similar to those of standard isolates of G. zeae phylogenetic lineage 7. All the populations are genetically similar, have high genotypic diversity and little or no detectable genetic disequilibrium, and show evidence of extensive
interpopulation genetic exchange. Allele frequencies in some of the populations examined are not statistically different from one another, but others are. The observed differences are relatively small, however, indicating that while genetic isolation by distance may occur, genetic exchange has occurred at a relatively high frequency among US populations of G. zeae. Gibberella zeae is the major fungal pathogen of Fusarium head blight of wheat and produces several mycotoxins harmful to humans and domesticated animals. We identified loci associated with pathogenicity and aggressiveness on an Amplified Fragment Length Polymorphism (AFLP)-based genetic map of G. zeae in a cross between a lineage 6 nivalenol producer from Japan and a lineage 7 deoxynivalenol producer from Kansas. Ninety-nine progeny and the parents were tested in the greenhouse in two years. Progeny segregated qualitatively 61:38 for pathogenicity:nonpathogenicity. The trait maps to linkage group IV adjacent to loci that
affect colony pigmentation, perithecium production, and trichothecene toxin amount. Among the 61 pathogenic progeny, the amount of disease induced (aggressiveness) varied quantitatively. Two reproducible quantitative trait loci (QTL) for aggressiveness were detected on linkage group I. A QTL linked to the TRI5 locus (trichodiene synthase in the trichothecene pathway gene cluster) explained 51% of the variation observed and a second QTL some 50 cM away, 29% of the phenotypic variation. TRI5 is tightly linked to the locus controlling trichothecene toxin type. Progeny that produced deoxynivalenol were, on average, about twice as aggressive as were those producing nivalenol. No transgressive segregation for aggressiveness was detected. The rather simple inheritance of both traits in this interlineage cross suggests that relatively few loci for pathogenicity or aggressiveness differ between lineage 6 and 7.
Impacts
All the populations of Gibberella zeae are genetically similar, have high genotypic diversity and little or no detectable genetic disequilibrium, and show evidence of extensive interpopulation genetic exchange. Allele frequencies in some of the populations examined are not statistically different from one another, but others are. Thus, the populations examined are not mere subsamples from a single, large, randomly mating population. Geographic distance and genetic distance between populations are correlated significantly. The observed differences are relatively small, however, indicating that while genetic isolation by distance may occur, genetic exchange has occurred at a relatively high frequency among US populations of G. zeae. These differences reflect the time required for the alleles to diffuse across the distances that separate them, because relatively little linkage disequilibrium is detected either in the population as a whole or in any of the individual
subpopulations.
Publications
- Summerell, B. A., and J. F. Leslie. 2004. Genetic diversity and population structure of plant pathogenic species in the genus Fusarium. In: Plant Microbiology (M. Gillings & A. Holmes, eds.), pp. 207-223. Bios, Oxford, United Kingdom. 290 pp.
- Cumagun, C. J. R., R. L. Bowden, J. E. Jurgenson, J. F. Leslie & T. Miedaner. 2004. Genetic mapping of pathogenicity and aggressiveness of Gibberella zeae (Fusarium graminearum) towards wheat. Phytopathology 94: 520-526.
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Progress 01/01/04 to 12/31/04
Outputs
Previous experiments have shown significant reduction on the level of fumonisin in the nixtamalized (cooked with 1.2% lime) corn samples when extruded at different temperatures, moisture contents, screw speeds and feed rates compared to the non-nixtamalized (cooked with water) samples. For this year, several experiments were done. 1) We tested more extrusion parameters such as die diameters and screw profiles again to find out their effects on the level of fumonisin after extruding the fumonisin contaminated non-nixtamalized and nixtamalized corn samples. Results showed that fumonisin was significantly reduced in the nixtamalized extrudates when compared to the non-nixtamalized extrudates. No significant effect was observed between different die diameters and screw profiles. 2) We also determine the biological toxicity of the fumonisin-contaminated non-nixtamalized and nixtamalized corn samples and corn extrudates to the brine shrimp (Artemia salina). We aimed to find
out if the amount of fumonisin remaining in the samples and/or the modified fumonisins (hydrolyzed fumonisins) that may have formed during the lime cooking and extrusion still possessed toxicity to biological organisms. Our results showed that the non-nixtamalized corn and corn extrudate samples, which had a higher concentration of fumonisin, were more toxic to brine shrimp larvae compared to the nixtamalixed corn and corn extrudate samples, which had lower concentrations of fumonisin. The hydrolyzed fumonisin, possibly formed in our nixtamalized samples, was not more toxic than the parent fumonisin. 3) The effect of moist heat (121oC for 20 min) and dry heat (150oC for 3 h) on fumonisin level was also tested in the non-fumonisin contaminated and fumonisin-contaminated non-nixtamalized and nixtamalized corn samples. Our results showed that the fumonisin reduction was higher in samples subjected to dry heat compared to moist heat. No fumonisin was detected after lime cooking the
non-fumonisin contaminated corn samples. Toxicity test of the samples by brine shrimp assay showed that fumonisin-contaminated corn samples that were nixtamalized were less toxic compared to the non-nixtamalized corn samples. Both the non-nixtamalized and nixtamalized corn samples that were not contaminated with fumonisin were not toxic to the brine shrimp larvae.
Impacts
The application of heat to food and feed materials has been reported by several authors to reduce fumonisin levels. We proved that heat treatment of the non-nixtamalized and nixtamalized corn by extrusion and moist and dry heat application reduced the level of the toxin. Since fumonisin is a major contaminant in corn worldwide, and a lot of foods are processed from corn by extrusion, cooking and baking, the knowledge about these processes in removing or minimizing the amount of fumonisins in the finished products is very important for human and animal health.
Publications
- No publications reported this period
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Progress 01/01/03 to 12/31/03
Outputs
Fumonisins are mycotoxins commonly found in corn and corn-based products that cause potential health hazards to humans and domestic animals. Corn samples were contaminated with fumonisin-producing Fusarium moniliforme 0488 and processed with lime (nixtamalized) and then extruded at 150oC and 175oC at various sample moisture contents ( 24%, 27%. 30% and 33%). Analysis of fumonisin by ELISA (Veratox for fumonisin), shows that nixtamalization significantly reduced the level of fumonisin in corn by 97% compared to 65% in non-nixtamalized corn. Lime treatment significantly lowered fumonisin levels by 75-100% after extrusion at 150oC and 175oC at the four moisture contents tested. The reduction of fumonisin in non-nixtamalized samples after extrusion ranged from 28-41%. However, temperature and moisture content had no significant effect on the level of fumonisin in the samples.
Impacts
Extrusion technology can offer a lot of possibilities to process sophisticated food products. Lime cooking can be used to eliminate hazardous substances like fumonisin. Since fumonisin is a major contaminant in corn worldwide, and a lot of foods are processed from corn by extrusion, the knowledge about extrusion being able to remove fumonisins in the finished products is very important, especially to corn processors and consumers.
Publications
- No publications reported this period
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Progress 01/01/02 to 12/31/02
Outputs
The focus of the research was to determine the effect of extrusion residence time on fumonisin in non-nixtamalized and nixtamalized corn samples. Fumonisin contaminated non-nixtamalized and nixtamalized corn samples, adjusted to 27 percent MC, were run in the extruder at low feed rate (16.7g/min) and high feed rate (37.4 g/min), and both at low screw speed (100 rpm) and high screw speed (200 rpm). Fumonisin concentrations in the non-extruded and extruded samples were analyzed by competitive direct ELISA (Veratox for fumonisin by Neogen). Results showed that extrusion can greatly decrease fumonisin contant, and the percent reduction was high in the nixtamalized samples than in the non-nixtamalized samples. At low feed rate, low screw speed, fumonisin was reduced by 72.1 percent in the non-nixtamalized samples and by 97.5 percent in the nixtamalized samples. At high screw speed, corresponding reductions were 78.6 percent and 98.3 percent, respectively. Also, at high
feed rate and low screw speed fumonisin was reduced by 76.2 percent in the non-nixtamalized samples and by 98.7 percent in the nixtamalized samples. At high screw speed, the reduction was 75.2 percent in the non-nixtamalized and 97.5 percent in the nixtamalized samples.
Impacts
Food processors must offer products that do not pose health hazards to the consumers. Mycotoxins if present, must be within the acceptable levels. Thermal processing has been shown to attain some of these objectives. Extrusion, a thermal food process technique, was shown to destroy fumonisin, a mycotoxin hazardous to man and animals. Extrusion cooking appears to reduce mycotoxin contamination food or feed products.
Publications
- Doyungan, S., Bh. Subramanyam and K. Behnke. 2002. Effect of extrusion temperature and sample moisture content on fumonisin in non-nixtamalized and nixtamalized corn. Journal of Stored-Products Research (In preparation).
- Doyungan, S., Bh. Subramanyam and S. Alavi. 2002. Influence of extrusion residence time on the reduction of fumonisin in non-nixtamalized and nixtamalized corn. Journal of Stored-Products Research (In preparation).
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Progress 01/01/01 to 12/31/01
Outputs
Effects of nixtamalization and extrusion on concentration of fumonisin in both Fusarium moniliforme 0488 inoculated and non-inoculated corn kernels were determined. The effect of varying extrusion temperatures and sample moisture contents on the concentration of the toxin were also quantified. Inoculated and non-inoculated corn kernels were boiled in distilled water (non-nixtamalized) and 1.2 percent Ca(OH)2 (nixtamalized), steeped for 16-18 h, washed, dried and ground in the Romer mill. Ground samples were adjusted to 24 percent, 27 percent, 30 percent and 33 percent moisture, and extruded at two temperatures, 150oC and 175oC. Results showed that nixtamalization reduced the amount of fumonisin by 97 percent in the F. moniliforme-inoculated samples. The remaining fumonisin amount was further reduced after extrusion at two temperatures at all moisture contents.
Impacts
Impact: One aspect of food safety is the control of mycotoxins like fumonisins in cereal products and the possibility of eliminating or reducing them to acceptable levels. Results of this study can be very useful to the tortilla manufacturing industry and makers of snack foods and breakfast cereals where corn products are extruded.
Publications
- No publications reported this period
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Progress 01/01/00 to 12/31/00
Outputs
This year's research was a continuation of a study on the effect of scarification (0, 10, and 20 sec) and chlorination (0, 200, 400, and 800 ppm) of wheat. In addition to determining deoxynivalenol (DON or vomitoxin), the level of microbial contamination in bran and flour samples were also determined. The colony forming units (cfu/g) of Escherischia coli were determined by plate count using 3M Petrifilm. The plates contain Violet Red Bile (VRB) nutrients which is an indicator of glucoronidase activity and tetrazolium indicator that facilitates colony enumeration. The presence of Salmonella sp. in the samples was determined using the Reveal Salmonella test detection system from Neogen. No E. coli was observed in flour and bran samples at scarification and chlorination levels indicated above. However, in a separate experiment using higher chlorine levels (1000, 5000 and 10000 ppm), 3 cfu/g E. coli were observed in one flour sample. Salmonella sp. was detected mostly in
the bran samples.
Impacts
Information on the effect of grain processing such as scarification and chlorination on the level of microorganisms in wheat will be very useful to grain processors, consumers, and regulators.
Publications
- Trigo-Stockli, D.M., Obaldo, L.G., Dominy, W.G., and Behnke, K.C. 2000. Utilization of deoxynivalenol-contaminated hard red winter wheat for shrimp feeds. Journal of the World Aquaculture Society 31(2):247-254.
- Reed, C.R., Trigo-Stockli, D.M., Blasi, D.A., and Fairchild, F.J. 2000. Storage of pelleted wheat middlings in farm bins. Animal Feed Science & Technology.
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Progress 01/01/99 to 12/31/99
Outputs
The focus of the research at this period was to determine the effects of scarification and chlorination on the level of deoxynivalenol (DON or vomitoxin) in wheat. Vomitoxin contamination appears to be a continuing problem with localized outbreak occurring almost annually. Vomitoxin causes feed refusal, vomiting, and immune suppression. The use of chlorine has been studied, however, chlorine levels at 0.1-0.5 percent did not cause a significant reduction of vomitoxin under normal milling and bleaching conditions. Physical methods such as cleaning and polishing reduce approximately 25 percent of vomitoxin but levels ranging from 60-80 percent remain in wheat flour. Scarification as a method to reduce the level of vomitoxin has not been studied previously. Our objective was to determine the combined effects of scarification and various concentrations of chlorine during tempering on the level of vomitoxin in milled fractions of wheat. Vomitoxin-contaminated soft white
wheat from Michigan was used in this study. A 1725 rpm/min scarifier,lined with a 40 grit sand paper, was used to scarify 500-g of wheat for 10 or 20 seconds. The level of calcium hypochlorite used were 0, 200, 400, and 800 ppm. Wheat was milled using a Brabender quadromat junior and vomitoxin on the bran and flour samples was determined using Gas Chromatography. Scarifying for 20 seconds caused a 24 percent reduction of vomitoxin in the bran. An 800 ppm chlorine also caused a 24 percent reduction of vomitoxin in the bran. Vomitoxin levels in the flour were similar among scarification and chlorine levels.
Impacts
Consumers are presently concerned about food safety. Information on the reduction of vomitoxin in milled products by scarification and chlorination will be very useful to processors, consumers, and regulators.
Publications
- TRIGO-STOCKLI, D.M., OBALDO, L.G., DOMINY, W.G., AND BEHNKE, K.C.(IN PRESS), 1999. UTILIZATION OF DEOXYNIVALENOL-CONTAMINATED HARD RED WINTER WHEAT FOR SHRIMP FEEDS. JOURNAL OF THE WORLD AQUACULTURE SOCIETY.
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Progress 01/01/98 to 12/31/98
Outputs
This year's research was a continuation of a study on the effect of high temperature and high pressure processing (extrusion) on fumonisin B1 (FB1) and hydrolyzed fumonisin B1 (HFB1) in corn. The objective was to determine whether pre-conditioning of samples before extrusion processing affect the level of FB1 and HFB1 in the extruded product. Fumonisin-contaminated and non-contaminated corn were nixtamalized using calcium hydroxide, ground, and dried. Before extrusion processing, in pre-conditioned samples the alkali-cooked flour was hydrated with water to reach the desired moistures of 24, 27, 30, and 33% using a Hobert blender and monostat pump. The pre-conditioned and non-conditioned samples were processed at 170 C in a Micro 18 extruder using an angular (3 mm) die. Fumonisin B1 and hydrolyzed fumonisin B1 were analyzed using reversed phase, C18 High performance liquid chromatography (HPLC). Results indicate that fumonisin levels were considerably reduced, however,
differences were not significant between pre-conditioned and non-conditioned treatments in terms of fumonisin recovery rate. The recoverable FB1 concentration of alkali-cooked corn was considerably affected by moisture content. In both the pre-conditioned and the non-conditioned treatments the reduction of the fumonisin content was higher in the 27-33% target moisture conditions. HFB1 was reduced less than FB1. It is possible that the low levels of fumonisins found in extruded samples may be due to decomposition during extrusion processing. It is also possible that the decomposition products from extrusion did not react with OPA reagent and thus were not detected by chromatography. A study on the effects of scarification and chlorination on the level of deoxynivalenol in wheat is being initiated.
Impacts
(N/A)
Publications
- TRIGO-STOCKLI, D.M., SANCHEZ-MARINEZ, R.I., CORTEZ-ROCHA, M.O. AND PEDERSEN, J.R. 1998. COMPARISON OF THE DISTRIBUTION AND OCCURRENCE OF FUSARIUM GRAMINEARUM AND DEOXYNIVALENOL IN HARD RED WINTER WHEAT FOR 1993-1996. CEREAL CHEM. 75:841-846.
- TRIGO-STOCKLI, D.M., DOMINY, W.G.,OBALDO, L.G.,TERPSTRA, J., CODY, J.J, AND BEHNKE, K.C. 1998. UTILIZATION OF DEOXYNIVALENOL-CONTAMINATED HARD RED WINTER WHEAT FOR SHRIMP FEEDS. CEREAL FOODS WORLD 43:548.
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Progress 01/01/97 to 12/31/97
Outputs
The focus of the research at this period was to determine the effect of high temperature and high pressure processing (extrusion) on fumonisin B1 (FB1) and hydrolyzed fumonisin B1 (HFB1) in corn. Hydrolyzed fumonisin B1 which is more toxic than the original FB1, is formed during alkali-cooking (nixtamalization) of corn to produce masa for tortillas and other corn products. Our survey of the literature indicated that thermally processed corn products (e.g. canned corn, tortillas, ready-to-eat cereals) generally have lower concentrations of fumonisins than unprocessed products (e.g. corn meal and grits). In this study, contaminated and noncontaminated corn were nixtamalized using calcium hydroxide, ground, and dried. The alkali-cooked flour was processed in an extruder using a tapered- angular (3 mm) and a tapered-circular (5 mm) die. Fumonisin B1 and hydrolyzed fumonisin B1 were analyzed using high performance liquid chromatography (HPLC). Our results indicate that
after nixtamalization, 61% of the original FB1 was present in the solid fraction while 39% was present in the steep water. Fumonisin B1 reductions of 99, 98, and 90% were obtained by extruding the alkali-cooked flour with a tapered-angular die at 24, 27, and 30% target processing moisture content, respectively. The greatest HFB1 reductions were 67 and 60% at 24 and 27% moisture content, respectively. With a tapered-circular die, the greatest reduction of FB1 (90%) and HFB1 (52%) occurred at 33% target moisture content.
Impacts
(N/A)
Publications
- CORTEZ-ROCHA, M.O., D.M. TRIGO-STOCKLI, D.L. WETZEL, and C.R. REED. 1997. Effect of Extrusion Processing on Hydrolyzed Fumonisin B1 in Corn. Cereal Foods World 42:621.
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Progress 01/01/96 to 12/30/96
Outputs
1996 Kansas hard red winter wheat quality survey samples were analyzed for Fusarium spp. & for vomitoxin (deoxynivalenol or DON). This 4th annual survey for grain fungi examined 179 samples collected from 85 of 105 Kansas counties. Highest levels of Fusarium infection (13.5 to 30%) occurred in 2 NE & 1 SE district counties. Avg. percent invasion of F. graminearum in samples for the State was 2.2%. Approx. 45% of the samples examined were free of Fusarium. Toxin analyses were made on 76 samples with varying levels of F. graminearum infection. TLC kits were used to determine ppm of DON using 1, 2 & 5 ppm standards. Of samples infected, 76.7% showed no DON. DON levels ranged from <0.5 to 12 ppm. Average levels >2 ppm were found in 2 counties in the NE district of the State. The NE district accounted for 4.7% of the total 1996 Kansas wheat production. Avg. level for the State was 0.47 ppm. Major wheat producing districts of the State generally had DON levels <1 ppm and
mostly zero. Samples with no invasion showed no toxins. Analysis of 1995 corn & sorghum samples from 15 locations across the State continues. Eight corn varieties from 10 locations represented in the 1995 survey were again sampled in 1996 & analyzed for Fusarium species & major Fusarium toxins. Emphasis was placed on fumonisins & DON. All corn analyzed from 1995 & 1996 samples contained F. moniliforme & some level of Fumonisin B1 (1.2-14 ppm).
Impacts
(N/A)
Publications
- TRIGO-STOCKLI, D.M., C.W. DEYOE, R. SATUMBAGA and J.R. PEDERSEN. 1996 Distribution of Deoxynivalenol and Zearalenone in Milled Fractions of Wheat. Cereal Chem. 73(3):388-391.
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Progress 01/01/95 to 12/30/95
Outputs
1995 Kansas hard red winter wheat quality survey samples were analyzed for Fusarium spp. & for vomitoxin (deoxynivalenol or DON). This 3rd survey for grain fungi examined 271 samples collected from 87 of 105 Kansas counties. Highest levels of Fusarium infection (32.7 to 67%) occurred in 4 counties of the NE, 2 of the NC, 1 of the EC & 1 of the SE districts. Average percent invasion of F. graminearum in samples for the State was 9.3%. Approximately 27% of the samples examined were free of Fusarium infection. Toxin analyses were made on 206 samples with varying levels of F. graminearum infection. TLC kits were used to determine parts per million (ppm) of DON using 1, 2 & 5 ppm standards. Of the samples infected, 76% showed no DON. Levels of DON ranged from <0.5 to >5 ppm. Average levels >2 ppm were found in 4 counties in the NE district of the State & 1 county each in the EC and SE districts. The NE district accounted for only 3% of the total 1995 Kansas wheat
production. The major wheat producing districts of the State generally had DON levels less than 1 ppm & mostly zero. Samples with no invasion showed no toxins. A major survey of corn & sorghum was initiated by obtaining samples from the 1995 Kansas Performance Tests conducted at 15 locations across the state. Tests included several varieties, hybrids & crosses produced under various agronomic systems & will provide an opportunity to compare these various factors in relationship to Fusarium infection & toxin production.
Impacts
(N/A)
Publications
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Progress 01/01/94 to 12/30/94
Outputs
1994 Kansas hard red winter wheat quality survey samples were analyzed for Fusarium spp., for vomitoxin (deoxynivalenol or DON) and for zearalenone. This 2nd survey for grain fungi examined 272 samples collected from 84 of 105 Kansas counties. Highest levels of infection (11, 13 and 36% invasion) occurred in 3 counties of the SE district which accounted for only 5.6% of the total 1994 Kansas wheat production. Average percent invasion of F. graminearum in samples for the State was 0.9%. In 1993, 4 counties in the NE corner of the State had infections which ranged as high as 75%. State average in 1993 was 2.4%. Differences reflect the contrast in weather conditions between the 2 years. Toxins analyses were made on 102 samples with varying levels of F. graminearum invasion. TLC kits were used to determine parts per million (ppm) of vomitoxin and zearalenone using 1, 2 and 5 ppm standards. Levels of vomitoxin ranged from <0.5 to 2.5 ppm and were found in the 3 counties
in the SE district of the State. Amount of vomitoxin found in sample(s) was directly related to the level of F graminearum invasion. Samples with no invasion showed no toxins. Samples of wheat (52) collected in 1993 were milled and flour and wheat by-products were analyzed for vomitoxin and zearalenone. When present, vomitoxin and zearalenone were highest in bran and shorts fractions and least in flour. Samples of 1994 crop sorghum and corn were also analyzed for Fusarium spp. and toxins.
Impacts
(N/A)
Publications
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Progress 01/01/93 to 12/30/93
Outputs
276 samples of wheat collected for the KS 1993 hard red winter wheat quality survey analyzed for the presence of Fusarium roseum (=graminearum) the fungus responsible for production of vomitoxin (deoxynivalenol or DON). Samples in varying numbers based on wheat production were collected from 89/105 KS counties. Percent invasion of seeds in samples from each county was determined. The avg. incidence of F.roseum for the State was 2.4% with the highest levels of infection occurring in the three Eastern districts which account for 7.1% of the total State wheat production. Four counties in the NE corner of the State had infections ranging from 0 to 75% invasion. Analyses for toxins were made on 116 samples with varying levels of F.roseum invasion. Romer Labs, Inc. TLC kits were used to determine parts per million (ppm) of vomitoxin, zearalenone, & aflatoxin by comparison with standards of 1, 2 and 5 ppm. Analyses indicated the highest levels of vomitoxin ranged from >2 to
>5 ppm found in the four counties in the NE corner of the State. In most cases the amount of vomitoxin found was directly related to the level of F.roseum contamination. Several samples showing no invasion were analyzed but none contained toxins. Approximately 50 samples of wheat were collected in sufficient quantity to allow milling. Flour and wheat by-products will also be analyzed for vomitoxin. Samples of grain sorghum from varietal trials in the State are in the process of being evaluated for F.roseum and toxins.
Impacts
(N/A)
Publications
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Progress 01/01/92 to 12/30/92
Outputs
A semi-field scale study using 0.5 ton lots of corn from 6 different fields determined whether a delay of up to 22 hours between harvest and application of a fungicide inhibited its ability to control mold development and its effect on aflatoxin production. Initial moisture content (m.c.) of corn ranged from 15 to 29 %. No significant difference in mold development was observed when fungicide application was delayed as much as 22 hrs. Freshness of diluted formulation did affect how well control of fungi growth. Solutions diluted for only 3 hrs lost potency. The fungicide treatment had no effect on the likelihood of aflatoxin production or the amount of toxin produced. No aflatoxin was found in corn samples immediately after harvest, and was produced only in those lots containing more than 20% m.c. A large scale trial in 3 (70 ton) lots compared untreated corn to corn treated at 15 and 20 ppm of the fungicide. Trends were analyzed and mean levels of dependent variables
were compared after a 22 to 25-day unheated air drying period. Grain m.c. at harvest ranged from 20 to 25%. The objective was to determine whether the fungicide would effectively limit the mold development and potential for aflatoxin production during the drying period. It required about 3 weeks to dry the corn to below 14% m.c. After drying, corn treated with 20 ppm contained about half the amount of Aspergillus invasion (17.8%) as the untreated and 15ppm lots (30.9 and 32.1%, respectively).
Impacts
(N/A)
Publications
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Progress 01/01/91 to 12/30/91
Outputs
The focus of the research during this period was to determine the effects of gamma irradiation on mycotoxins in grain with different moisture contents. Triplicate samples of wheat, corn, and soybeans containing 9, 13, or 17% moisture were inoculated with 1000 ppb aflatoxin B(subscript 1), then exposed to doses of 5, 7.5, 10, or 20 kGy of gamma irradiation from a cobalt-60 source. Deoxynivalenol (10 ppm) was added to soybeans, T-2 toxin (5 ppm) to wheat, and zearalenone (5 ppm) to corn, moisture in the grains was adjusted as above, and the grains were similarly exposed to gamma irradiation. Ten kGy of gamma irradiation significantly reduced zearalenone in corn, deoxynivalenol in soybeans, and T-2 toxin in wheat. It appeared that aflatoxin added to corn and wheat was not efficiently extracted by the Neogen Agri-Screen kits used for the assays, and no irradiation effect was measurable in those samples. A small, but significant, reduction in aflatoxin in soybeans was noted
with 10 kGy irradiation. Moisture contents of the samples had no significant effect on toxin stability.
Impacts
(N/A)
Publications
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Progress 01/01/90 to 12/30/90
Outputs
Whereas previous work focused on the effects of gamma irradiation on mycotoxin-producing fungi in grains and legumes, our current focus it to examine the effects of ionizing radiation on mycotoxins. Few studies have been reported, and results are conflicting. Some work indicates the toxins are quite susceptible to irradiation, but other studies show little effect. The study currently in progress will determine the effects of gamma irradiation on aflatoxin B1, T-2 toxin, zearelenone, and vomitoxin in corn, wheat, and soybeans at different moisture levels. Results to date indicate that irradiation doses up to 20 KGy are not effective at reducing aflatoxin B1 concentration in wheat, corn, or soybeans at 9, 13 or 17% moisture levels. In addition, we are in the process of determining the effects of gamma irradiation on the protein quality of the grains.
Impacts
(N/A)
Publications
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Progress 01/01/89 to 12/30/89
Outputs
Research on the effects of gamma irradiation dose on field and storage fungi in wheat, maize, and soybeans was continued. With increasing radiation dose rates (from 1-10 kGy) there was a substantial decrease in the number of surface fungi for all cultivars tested. Soybeans showed no viable propagules after only 2,5 kGy of irradiation. Some viable propagules remained in corn and wheat after 5 kGy irradiation, but none were found after the 10 kGy dose. Storage fungi were more susceptible to irradiation than field fungi on grain surfaces. As previously noted, no fungi survived the 10 kGy irradiation dose. Alternaria, fusarium, cephalosporium, and rhizopus all survived 5 kGy irratiation. A. Candidus, A. flavus, A. glaucus, penicillium citrinum, and A. niger survived the 2.5 kGy dose. Irradiated seeds appeared to be more susceptible to invasion by common storage fungi than non-irradiated seeds, but when the experiment was repeated that was not found to be the case.
Susceptibility of irradiated seeds to fungal invasion needs further evaluation.
Impacts
(N/A)
Publications
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Progress 01/01/88 to 12/30/88
Outputs
Wheat, maize, and soybean seeds were inoculated with A. candidus, A. restrictus,A. repens and Penicillium citrinum. Portions of the inoculated samples were the irradiated with a cobalt-60 gamma cell at dose rates of 0, 1, 2.5, 5, or 10 kGy. The survival of field and storage fungi on the seed surfaces was measured by the dilution count technique. Samples of the wheat, maize, and soybean seeds were then hydrated to 15 or 17% moisture content, then inoculated with storage fungi as above and stored for 30 days. After air-drying, samples were then irradiated as before. After disinfecting the seed surface with bleach solution 100 seed from each treatment were planted in malt agar media. Percent of seeds invaded by field or storage fungi was noted after 5 days. Data showed that there was a substantial decrease in the number propagules per gram for all cultivars with increasing radiation dose. In soybeans all fungi were eliminated at 2.5kGy, and at a dose rate of 5kGy
no count was observed in maize. In wheat the fungi survived up to 5kGy. At a dose rate of 10 kGy all the fungi in all cultivars were killed. Field fungi were more susceptible to radiation than storage fungi.
Impacts
(N/A)
Publications
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Progress 01/01/87 to 12/30/87
Outputs
Outline of experiment to be performed in April, 1987: Objective: To determine the effect of irradiation dose on mold population and mycotoxin formation in stored cereals and legumes. Rationale: Chemical treatments to control microbiological infestations in stored agricultural products may have adverse effects on human and animal health. Irradiation offers a "no residue" alternative, but, at present, little is known about the effects of radiation treatment on the microbiological status of stored grains and legumes. Procedure: Samples of wheat, corn, and soybeans will be irradiated at 0, 50, 250, or 500 kilorads. Mold population and mycotoxin levels will then be measured in the control and irradiated samples.
Impacts
(N/A)
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Progress 01/01/86 to 12/30/86
Outputs
Research continued to investigate the ability of the non-tannin polyphenols of Sorghum (Sorghum bicolor (L.) Moench) to inhibit the growth of mycotoxin producing fungi. The four toxin-producing fungi previously utilized for in vitro studies (two Fusaria and two Alternaria) were again used in all experiments. A series of experiments compared (in vitro) the effects of the free versus bound phenolic acids present in types I and III sorghum grain. The previously bound acids were more potent inhibitors of fungal growth than were the free (unesterified) compounds. This difference in growth inhibitory activity was shown for all fungal isolates tested although the extent in the difference depended on the isolate. Since extraction of the bound acids from the grain involved acidic hydrolysis, the possibility existed that oxidative changes were taking place in the compounds and that these changes resulted in increased inhibitory activity. This was tested by subjecting free
acid extracts to the conditions encountered during isolation of the bound compounds. When treated in this manner, the growth inhibitory potency of these extracts was now equivalent to the bound acid extracts, supporting the above hypothesis. Oxidation of a mixture of three model phenolic acids vis acid treatment also resulted in increases in their ability to inhibit fungal growth in vitro.
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Progress 01/01/85 to 12/30/85
Outputs
Over the past 12 months, research has continued to study the non-tannin polyphenols of sorghum (Sorghum bicolor (L.) Moench) and their ability to inhibit the growth and/or toxin production of mycotoxin producing fungi. All studies have utilized the four toxin-producing fungi (two Alternaria and two Fusaria) previously isolated. Extracts of the free and esterified phenolic acids from High (Type III) and Low tannin (Types II and II) sorghums were tested, in vitro for their ability to inhibit fungal growth. All extracts exhibited inhibitory activity against all four fungi tested. In all cases, (ie., at each extract concentration), Fusaria species showed greater reductions in growth than did Alternaria. When incorporated into nutrient agar as equivalent percentages of the phenolic acids extracted from constant amounts of grain, extracts of type III sorghum proved to be more inhibitory to fungal growth than did extracts from types I and II grain. Subsequent analyses
demonstrated that both type III grain and its extracts contained a higher concentration of phenolic acids than did types I and II. When tested in vitro on the basis of equivalent total phenol concentration, no differences in inhibitory activity were found. Experiments are underway to determine which of the 8 phenolic acids, found in the extracts exert the observed growth inhibition, singly or in combination.
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Progress 01/01/84 to 12/30/84
Outputs
In the past 12 months, research efforts have begun to study the non-tannin polyphenols of sorghum (Sorghum bicolor, L. Moench) and their ability to inhibit the growth and/or toxin production of mycotoxin producing fungi. Four toxin producing fungi (two Alternaria and two Fusaria) have been isolated from Kansas-grown sorghum and corn. Laboratory methods were developed to assess the rate of in vitro mycelial growth and spore germination. Results demonstrated that all four fungi were identifiabily different in their rate of germination. Differences in mycelial growth rates were not seen between types of the same genus but were seen between genera. Free and bound non-tannin polyphenols were extracted from high (Type III) and low tannin (Types I and II) sorghums. HPLC analyses of the extracts indicated that they differed in both the amount and types of cinnamic and benzoic acids they contained. In vitro studies using purified samples of the three most prevalent acids
(ferulic, p-coumaric and p-hydroxygenzoic) showed that all were capable of inhibiting the mycelial growth of all four fungi. Tests are underway to assess that ability of the extracts to inhibit spore germination and in vitro toxin production.
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Progress 01/01/83 to 12/30/83
Outputs
Studies were initiated to determine the effects of the use of mycotoxin contaminated grain on the whole stillage resulting from alcohol production. Aflatoxin contaminated corn and DON contaminated wheat were used as feedstocks. With either feed stock, toxin was not detected in the resulting alcohol. However, both toxins were effectively concentrated in the by-product stillage. This problem will have to be addressed before such stillage could be used as a feed ingredient. Nutrient analysis of the stillage showed no compositional differences due to treatment (ie infection). There were some differences in amino acid composition, slight decreases in glutamic acid and half cystine and increases in alanine, methionine and isoleucine. Analyses of Kansas wheat quality profile samples for DON indicated that at least 80% and, perhaps as much as 90% of the 1982 Kansas crop was not scabby. Studies incorporating vomitoxin containing wheat in starter pig rations suggested that
when toxin levels exceed 1 ppm, a reduction in gain and feed intake can be expected. Feed refusal will be common but no noticable sickness will be observed. Similar tests on the effects of scabby wheat in growing and finishing rations suggested that feed effeciency and weight gain decreased slightly when 25% or 50% of the diet's sorghum grain was replaced by scabby wheat. Higher percentages of substitution reduced feed intake drastically.
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Progress 01/01/82 to 12/30/82
Outputs
Studies are continuing to examine the effect of mycotoxin contamination on the alcohol fermentation process. Mycotoxins have been produced on both wheat and grain sorghum and the grain used in the production of ethanol. Of primary interest is the fate of the mycotoxin left in the stillage. Several studies and surveys have been conducted on wheat to determine the extent of vomitoxin (deoxynivalenol) contamination in the 1982 Kansas crop. A state-wide survey of 50 elevators indicated that the problem was confined to the N.E. part of the state. Cleaning, milling and baking studies are being conducted with wheat known to contain various levels of vomitoxin.
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Progress 01/01/81 to 12/30/81
Outputs
A trial has been initiated using a pilot scale fermentation system to examine the effects of myco-toxin contamination from A. flavors on the fermentation process. Mycotoxins produced on both wheat and grain sorghum will be studied. Both sound and contaminated grain from the same lot will be used to determine negative effects on the enzymes and/or yeast and to elucidate, quantitatively, the fate of the myco-toxin. High pressure liquid chromatography will be used to quantify the myco-toxin levels in all products and by-products.
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Progress 01/01/80 to 12/30/80
Outputs
Initial screening of 29 grain dust samples from Kansas and Nebraska gave no positives for aflatoxin because the 1979 crop was essentially free of A. flavus. Alternaria metabolites in sorghum grain dust confirmed contentions that dust does reflect parental grain composition however. Dust from moldy corn should not be used for feed unless demonstrated to be free of aflatoxin. Technical assistance and sampling procedures were offered to all feed processors in Kansas. No processors requested assistance due to a lack of aflatoxin incidence. Turkey poults were used to evaluate several processing and chemical treatment methods. Heat and pressure in combination with chemicals (NA(2)CO(3), NaOH or NH(3)) was necessary to effectively detoxify aflatoxin. Detoxification by chemical means alone were apparently reversible under conditions in the turkey poult gut.
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Progress 01/01/79 to 12/30/79
Outputs
This project is in the early developmental phase with no data accumulated yet. Field incidence of Aspergillus flavus in Kansas was negligible this past summer and no systematic sampling of freshly binned corn was necessary. Grain bin sampling will begin with the advent of warmer weather. Grain dust samples have been collected and some preliminary analyses completed.
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