Food Safety Aspects of Breeding Maize to Multi-Resistance against the Major (Fusarium graminearum, F. verticillioides, Aspergillus flavus) and Minor Toxigenic Fungi (Fusarium spp.) as Well as to Toxin Accumulation, Trends, and Solutions-A Review.

Maize is the crop which is most commonly exposed to toxigenic fungi that produce many toxins that are harmful to humans and animals alike. Preharvest grain yield loss, preharvest toxin contamination (at harvest), and storage loss are estimated to be between 220 and 265 million metric tons. In the past ten years, the preharvest mycotoxin damage was stable or increased mainly in aflatoxin and fumonisins. The presence of multiple toxins is characteristic. The few breeding programs concentrate on one of the three main toxigenic fungi. About 90% of the experiments except AFB1 rarely test toxin contamination. As disease resistance and resistance to toxin contamination often differ in regard to F. graminearum, F. verticillioides, and A. flavus and their toxins, it is not possible to make a food safety evaluation according to symptom severity alone. The inheritance of the resistance is polygenic, often mixed with epistatic and additive effects, but only a minor part of their phenotypic variation can be explained. All tests are made by a single inoculum (pure isolate or mixture). Genotype ranking differs between isolates and according to aggressiveness level; therefore, the reliability of such resistance data is often problematic. Silk channel inoculation often causes lower ear rot severity than we find in kernel resistance tests. These explain the slow progress and raise skepticism towards resistance breeding. On the other hand, during genetic research, several effective putative resistance genes were identified, and some overlapped with known QTLs. QTLs were identified as securing specific or general resistance to different toxicogenic species. Hybrids were identified with good disease and toxin resistance to the three toxigenic species. Resistance and toxin differences were often tenfold or higher, allowing for the introduction of the resistance and resistance to toxin accumulation tests in the variety testing and the evaluation of the food safety risks of the hybrids within 2-3 years. Beyond this, resistance breeding programs and genetic investigations (QTL-analyses, GWAM tests, etc.) can be improved. All other research may use it with success, where artificial inoculation is necessary. The multi-toxin data reveal more toxins than we can treat now. Their control is not solved. As limits for nonregulated toxins can be introduced, or the existing regulations can be made to be stricter, the research should start. We should mention that a higher resistance to F. verticillioides and A. flavus can be very useful to balance the detrimental effect of hotter and dryer seasons on aflatoxin and fumonisin contamination. This is a new aspect to secure food and feed safety under otherwise damaging climatic conditions. The more resistant hybrids are to the three main agents, the more likely we are to reduce the toxin losses mentioned by about 50% or higher.

What Is Fusarium Head Blight (FHB) Resistance and What Are Its Food Safety Risks in Wheat? Problems and Solutions-A Review.

The term "Fusarium Head Blight" (FHB) resistance supposedly covers common resistances to different Fusarium spp. without any generally accepted evidence. For food safety, all should be considered with their toxins, except for deoxynivalenol (DON). Disease index (DI), scabby kernels (FDK), and DON steadily result from FHB, and even the genetic regulation of Fusarium spp. may differ; therefore, multitoxin contamination is common. The resistance types of FHB form a rather complex syndrome that has been the subject of debate for decades. It seems that resistance types are not independent variables but rather a series of components that follow disease and epidemic development; their genetic regulation may differ. Spraying inoculation (Type 1 resistance) includes the phase where spores land on palea and lemma and spread to the ovarium and also includes the spread-inhibiting resistance factor; therefore, it provides the overall resistance that is needed. A significant part of Type 1-resistant QTLs could, therefore, be Type 2, requiring the retesting of the QTLs; this is, at least, the case for the most effective ones. The updated resistance components are as follows: Component 1 is overall resistance, as discussed above; Component 2 includes spreading from the ovarium through the head, which is a part of Component 1; Component 3 includes factors from grain development to ripening (FDK); Component 4 includes factors influencing DON contamination, decrease, overproduction, and relative toxin resistance; and for Component 5, the tolerance has a low significance without new results. Independent QTLs with different functions can be identified for one or more traits. Resistance to different Fusarium spp. seems to be connected; it is species non-specific, but further research is necessary. Their toxin relations are unknown. DI, FDK, and DON should be checked as they serve as the basic data for the risk analysis of cultivars. A better understanding of the multitoxin risk is needed regarding resistance to the main Fusarium spp.; therefore, an updated testing methodology is suggested. This will provide more precise data for research, genetics, and variety registration. In winter and spring wheat, the existing resistance level is very high, close to Sumai 3, and provides much greater food safety combined with sophisticated fungicide preventive control and other practices in commercial production.

The Role of Preharvest Natural Infection and Toxin Contamination in Food and Feed Safety in Maize, South-East Hungary, 2014-2021.

Mycotoxins originating in the preharvest period represent a less studied research problem, even though they are of the utmost practical significance in maize production, determining marketability (within EU limits), and storage ability, competitiveness, and profit rate. In this study, 18-23 commercial hybrids were tested between 2014 and 2021. Natural infection from Fusarium spp. was higher than 1.5%, and for Aspergillus spp. this was normally 0.01% or 0, much lower than would be considered as severe infection. In spite of this, many hybrids provided far higher toxin contamination than regulations allow. The maximum preharvest aflatoxin B1 was in 2020 (at 2286 µg/kg), and, in several cases, the value was higher than 1000 µg/kg. The hybrid differences were large. In Hungary, the presence of field-originated aflatoxin B1 was continuous, with three AFB1 epidemics in the 8 years. The highest DON contamination was in 2014 (at 27 mg/kg), and a detectable DON level was found in every hybrid. FUMB1+B2 were the highest in 2014 (at 45.78 mg/kg). At these low infection levels, correlations between visual symptoms and toxin contaminations were mostly non-significant, so it is not feasible to draw a conclusion about toxin contamination from ear rot coverage alone. The toxin contamination of hybrids for a percentage of visual infection is highly variable, and only toxin data can decide about food safety. Hybrids with no visual symptoms and high AFB1 contamination were also identified. Preharvest control, including breeding and variety registration, is therefore of the utmost importance to all three pathogens. Even natural ear rot and toxin data do not prove differences in resistance, so a high ear rot or toxin contamination level should be considered as a risk factor for hybrids. The toxin control of freshly harvested grain is vital for separating healthy and contaminated lots. In addition, proper growing and storage conditions must be ensured to protect the feed safety of the grain.

Novel Insights into the Inheritance of Gibberella Ear Rot (GER), Deoxynivalenol (DON) Accumulation, and DON Production.

Gibberella ear rot (GER) is an important fungal ear pathogen of maize that causes ear rot and toxin contamination. Most previous works have only dealt with the visual symptoms, but not with the toxins of GER. As food and feed safety rankings depend on toxin contamination, including deoxynivalenol (DON), without toxins, nothing can be said about the risks involved in food and feed quality. Therefore, three susceptible, three medium-susceptible, and three medium-resistant mother lines were crossed with three testers with differing degrees of resistance and tested between 2017-2020. Two plot replicates and two fungal strains were used separately. The highest heterosis was found at the GER% with a 13% increase across 27 hybrids, including 7 hybrids showing negative heterosis (a higher hybrid performance above the parental mean), with a variance ranging between 63.5 and -55.4. For DON, the mean heterosis was negative at -35%, and only 10 of the 27 hybrids showed a positive heterosis. The mean heterosis for DON contamination, at 1% GER, was again negative (-19.6%, varying between 85% and 224%). Only 17 hybrids showed heterosis, while that of the other 17 was rated higher than the parental mean. A positive significant correlation was found only for GER% and DON; the other factors were not significant. Seven hybrids were identified with positive (2) or negative (5) heterosis for all traits, while the rest varied. For DON and GER, only 13 provided identical (positive or negative) heteroses. The majority of the hybrids appeared to diverge in the regulation of the three traits. The stability of GER and DON (variance across eight data sets) did not agree-only half of the genotypes responded similarly for the two traits. The genetic background for this trait is unknown, and there was no general agreement between traits. Thus, without toxin analyses, the evaluation of food safety is not possible. The variety in degrees of resistance to toxigenic fungi and resistance to toxin accumulation is an inevitable factor.

Updating the Methodology of Identifying Maize Hybrids Resistant to Ear Rot Pathogens and Their Toxins-Artificial Inoculation Tests for Kernel Resistance to Fusarium graminearum, F. verticillioides, and Aspergillus flavus.

Resistance to toxigenic fungi and their toxins in maize is a highly important research topic, as mean global losses are estimated at about 10% of the yield. Resistance and toxin data of the hybrids are mostly not given, so farmers are not informed about the food safety risks of their grown hybrids. According to the findings aflatoxin regularly occurs at preharvest in Hungary and possibly other countries in the region can be jeopardized. We tested, with an improved methodology (two isolates, three pathogens, and a toxin control), 18 commercial hybrids (2017-2020) for kernel resistance (%), and for toxin contamination separately by two-two isolates of F. graminearum, F. verticillioides (mg/kg), and A. flavus (µg/kg). The preharvest toxin contamination was measured in the controls. Highly significant kernel resistance and toxin content differences were identified between hybrids to the different fungi. Extreme high toxin production was found for each toxic species. Only about 10-15% of the hybrids showed higher resistance to the fungal species tested and lower contamination level of their toxins. The lacking correlations between resistance to different fungi and toxins suggest that resistance to different fungi and response to toxin contamination inherits independently, so a toxin analysis is necessary. For safety risk estimation, separated artificial and natural kernel infection and toxin data are needed against all pathogens. Higher resistance to A. flavus and F. verticillioides stabilizes or improves feed safety in hot and dry summers, balancing the harmful effect of climate changes. Resistance and toxin tests during variety registration is an utmost necessity. The exclusion of susceptible or highly susceptible hybrids from commercial production results in reduced toxin contamination.

Key Global Actions for Mycotoxin Management in Wheat and Other Small Grains.

Mycotoxins in small grains are a significant and long-standing problem. These contaminants may be produced by members of several fungal genera, including Alternaria, Aspergillus, Fusarium, Claviceps, and Penicillium. Interventions that limit contamination can be made both pre-harvest and post-harvest. Many problems and strategies to control them and the toxins they produce are similar regardless of the location at which they are employed, while others are more common in some areas than in others. Increased knowledge of host-plant resistance, better agronomic methods, improved fungicide management, and better storage strategies all have application on a global basis. We summarize the major pre- and post-harvest control strategies currently in use. In the area of pre-harvest, these include resistant host lines, fungicides and their application guided by epidemiological models, and multiple cultural practices. In the area of post-harvest, drying, storage, cleaning and sorting, and some end-product processes were the most important at the global level. We also employed the Nominal Group discussion technique to identify and prioritize potential steps forward and to reduce problems associated with human and animal consumption of these grains. Identifying existing and potentially novel mechanisms to effectively manage mycotoxin problems in these grains is essential to ensure the safety of humans and domesticated animals that consume these grains.

Perspectives on Global Mycotoxin Issues and Management From the MycoKey Maize Working Group.

During the last decade, there have been many advances in research and technology that have greatly contributed to expanded capabilities and knowledge in detection and measurement, characterization, biosynthesis, and management of mycotoxins in maize. MycoKey, an EU-funded Horizon 2020 project, was established to advance knowledge and technology transfer around the globe to address mycotoxin impacts in key food and feed chains. MycoKey included several working groups comprising international experts in different fields of mycotoxicology. The MycoKey Maize Working Group recently convened to gather information and strategize for the development and implementation of solutions to the maize mycotoxin problem in light of current and emerging technologies. This feature summarizes the Maize WG discussion and recommendations for addressing mycotoxin problems in maize. Discussions focused on aflatoxins, deoxynivalenol, fumonisins, and zearalenone, which are the most widespread and persistently important mycotoxins in maize. Although regional differences were recognized, there was consensus about many of the priorities for research and effective management strategies. For preharvest management, genetic resistance and selecting adapted maize genotypes, along with insect management, were among the most fruitful strategies identified across the mycotoxin groups. For postharvest management, the most important practices included timely harvest, rapid grain drying, grain cleaning, and carefully managed storage conditions. Remediation practices such as optical sorting, density separation, milling, and chemical detoxification were also suggested. Future research and communication priorities included advanced breeding technologies, development of risk assessment tools, and the development and dissemination of regionally relevant management guidelines.

Updating the Breeding Philosophy of Wheat to Fusarium Head Blight (FHB): Resistance Components, QTL Identification, and Phenotyping-A Review.

Fusarium head blight has posed continuous risks to wheat production worldwide due to its effects on yield, and the fungus provides additional risks with production of toxins. Plant resistance is thought to be the most powerful method. The host plant resistance is complex, Types I-V were reported. From the time of spraying inoculation (Type I), all resistance types can be identified and used to determine the total resistance. Type II resistance (at point inoculation) describes the spread of head blight from the ovary to the other parts of the head. Therefore, it cannot solve the resistance problem alone. Type II QTL (quantitative trait locus) Fhb1 on 3BS from Sumai 3 descendant CM82036 secures about the same resistance level as Type I QTL does on 5AS and 5ASc in terms of visual symptoms, FDK (Fusarium damaged kernel), and deoxynivalenol response. Recently, increasing evidence supports the association of deoxynivalenol (DON) content and low kernel infection with FHB (Fusarium head blight) resistance (Types III and IV), as QTL for individual resistance types has been identified. In plant breeding practice, the role of visual selection remains vital, but the higher correlations for FDK/DON make it possible to select low-DON genotypes via FDK value. For phenotyping, the use of more independent inocula (isolates or mixtures) makes resistance evaluation more reliable. The large heterogeneity of the mapping populations is a serious source of underestimating genetic effects. Therefore, the increasing of homogeneity is a necessity. As no wheat varieties exist with full resistance to FHB, crops must be supported by proper agronomy and fungicide use.

Methodical Considerations and Resistance Evaluation against F. graminearum and F. culmorum Head Blight in Wheat. The Influence of Mixture of Isolates on Aggressiveness and Resistance Expression.

In resistance tests to Fusarium head blight (FHB), the mixing of inocula before inoculation is normal, but no information about the background of mixing was given. Therefore, four experiments (2013-2015) were made with four independent isolates, their all-possible (11) mixtures and a control. Four cultivars with differing FHB resistance were used. Disease index (DI), Fusarium damaged kernels (FDK) and deoxynivalenol (DON) were evaluated. The isolates used were not stable in aggressiveness. Their mixtures did not also give a stable aggressiveness; it depended on the composition of mix. The three traits diverged in their responses. After the mixing, the aggressiveness was always less than that of the most pathogenic component was. However, in most cases it was significantly higher than the arithmetical mean of the participating isolates. A mixture was not better than a single isolate was. The prediction of the aggressiveness level is problematic even if the aggressiveness of the components was tested. Resistance expression is different in the mixing variants and in the three traits tested. Of them, DON is the most sensitive. More reliable resistance and toxin data can be received when instead of one more independent isolates are used. This is important when highly correct data are needed (genetic research or cultivar registration).

The Influence of the Dilution Rate on the Aggressiveness of Inocula and the Expression of Resistance against Fusarium Head Blight in Wheat.

In previous research, conidium concentrations varying between 10,000 and 1,000,000/mL have not been related to any aggressiveness test. Therefore, two Fusarium graminearum and two Fusarium culmorum isolates were tested in the field on seven genotypes highly differing in resistance at no dilution, and 1:1, 1:2, 1:4, 1:8, and 1:16 dilutions in two years (2013 and 2014). The isolates showed different aggressiveness, which changed significantly at different dilution rates for disease index (DI), Fusarium-damaged kernels (FDK), and deoxynivalenol (DON). The traits also had diverging responses to the infection. The effect of the dilution could not be forecasted. The genotype ranks also varied. Dilution seldomly increased aggressiveness, but often lower aggressiveness occurred at high variation. The maximum and minimum values varied between 15% and 40% for traits and dilutions. The reductions between the non-diluted and diluted values (total means) for DI ranged from 6% and 33%, for FDK 8.3-37.7%, and for DON 5.8-44.8%. The most sensitive and most important trait was DON. The introduction of the aggressiveness test provides improved regulation compared to the uncontrolled manipulation of the conidium concentration. The use of more isolates significantly increases the credibility of phenotyping in genetic and cultivar registration studies.

Methodical Considerations and Resistance Evaluation Against Fusarium graminearum and F. culmorum Head Blight in Wheat. Part 3. Susceptibility Window and Resistance Expression.

Flowering is the most favorable host stage for Fusarium infection in wheat, which is called the susceptibility window (SW). It is not known how long it takes, how it changes in different resistance classes, nor how stable is the plant reaction in the SW. We have no information, how the traits disease index (DI), Fusarium-damaged kernel rate (FDK), and deoxynivalenol (DON) respond within the 16 days period. Seven winter wheat genotypes differing in resistance were tested (2013-2014). Four Fusarium isolates were used for inoculation at mid-anthesis, and 4, 8, 11, 13, and 16 days thereafter. The DI was not suitable to determine the length of the SW. In the Fusarium-damaged kernels (FDK), a sharp 50% decrease was found after the 8th day. The largest reduction (above 60%) was recorded for DON at each resistance level between the 8th and 11th day. This trait showed the SW most precisely. The SW is reasonably stable in the first 8-9 days. This fits for all resistance classes. The use of four isolates significantly improved the reliability and credit of the testing. The stable eight-day long SW helps to reduce the number of inoculations. The most important trait to determine the SW is the DON reaction and not the visual symptoms.

A New Concept to Secure Food Safety Standards against Fusarium Species and Aspergillus Flavus and Their Toxins in Maize.

Commercial maize hybrids are exposed to different degrees of ear infection by toxigenic fungal species and toxin contamination. Their resistance to different fungi and toxin relationships are largely unknown. Without this knowledge, screening and breeding are not possible for these pathogens. Seven- to tenfold differences were found in resistance to Fusarium spp., and there was a five-fold difference in ear coverage (%) in response to A. flavus. Three hybrids of the twenty entries had lower infection severity compared with the general means for toxigenic species. Three were highly susceptible to each, and 14 hybrids reacted differently to the different fungi. Differences were also observed in the toxin content. Again, three hybrids had lower toxin content in response to all toxigenic species, one had higher values for all, and 16 had variable resistance levels. Correlations between infection severity and deoxynivalenol (DON) content were 0.95 and 0.82 (p = 0.001) for F. graminearum and F. culmorum, respectively. For fumonisin and F. verticillioides ear rot, the Pearson correlation coefficient (r) was 0.45 (p = 0.05). Two independent isolates with different aggressiveness were used, and their mean X values better described the resistance levels. This increased the reliability of the data. With the introduction of this methodological concept (testing the resistance levels separately for different fungi and with two isolates independently), highly significant resistance differences were found. The resistance to different fungal species correlated only in certain cases; thus, each should be tested separately. This is very useful in registration tests and post-registration screening and breeding. This would allow a rapid increase in food and feed safety.

Advocating a need for suitable breeding approaches to boost integrated pest management: a European perspective.

Currently, European farmers do not have access to sufficient numbers and diversity of crop species/varieties. This prevents them from designing cropping systems more resilient to abiotic and biotic stresses. Crop diversification is a key lever to reduce pest (pathogens, animal pests and weeds) pressures at all spatial levels from fields to landscapes. In this context, plant breeding should consist of: (1) increased efforts in the development of new or minor crop varieties to foster diversity in cropping systems, and (2) focus on more resilient varieties showing local adaptation. This new breeding paradigm, called here 'breeding for integrated pest management (IPM)', may boost IPM through the development of cultivars with tolerance or resistance to key pests, with the goal of reducing reliance on conventional pesticides. At the same time, this paradigm has legal and practical implications for future breeding programs, including those targeting sustainable agricultural systems. By putting these issues into the context, this article presents the key outcomes of a questionnaire survey and experts' views expressed during an EU workshop entitled 'Breeding for IPM in sustainable agricultural systems'. © 2017 Society of Chemical Industry.

Distribution of prothioconazole and tebuconazole between wheat ears and flag leaves following fungicide spraying with different nozzle types at flowering.

BACKGROUND: Wheat ears are difficult targets from the aspect of fungicide spraying. Sideward-spraying nozzle types may enhance the ear coverage, which may possibly lead to higher effectiveness in the management of Fusarium head blight (FHB). RESULTS: On average, sideward-spraying Turbo TeeJet Duo nozzles resulted in 1.30 and 1.43 times higher prothioconazole-desthio and tebuconazole contents and Turbo FloodJet nozzles in 1.08 and 1.34 times higher prothioconazole-desthio and tebuconazole contents in wheat ears by comparison with those achieved with vertically-spraying XR TeeJet nozzles. In contrast, the vertically-spraying XR TeeJet nozzles resulted in 1.57 and 1.31 times higher prothioconazole-desthio and tebuconazole contents in the flag leaf blade. The degradation of the active ingredient (AI) depended on the year, the cultivar and the plant organ, but not on the spraying method. There was no clear relationship between the efficacy of a given nozzle type and the outcome of the FHB epidemic. CONCLUSIONS: The ear coverage and therefore the AI content have been improved with the two sideward-spraying nozzle types. There was no effective translocation of the AI content between the ears and flag leaf blades. Prothioconazole and tebuconazole proved to be highly effective in the management of FHB, but the FHB resistance of the cultivar was also decisive.

Detection of previously unknown fumonisin P analogue mycotoxins in a Fusarium verticillioides culture by high-performance liquid chromatography-electrospray ionization time-of-flight and ion trap mass spectrometry.

Five previously unknown fumonisin mycotoxins (iso-FP1, iso-FP(2,3a), iso-FP(2,3b), FP4 and iso-FP4) and three previously described FP analogues (FP(1-3)) were detected in a solid rice culture inoculated with Fusarium verticillioides. The fumonisins were characterized by high-performance liquid chromatography-electrospray ionization time-of-flight mass spectrometry (HPLC-ESI-TOFMS) and ion trap mass spectrometry (ITMS). The FP isomers were separated by using a flat gradient on a special, high-coverage C18, narrow-bore HPLC column (YMC-Pack J'sphere ODS H80), which was suggested for the separation of structural isomers. The verified structures of the FP(1-3) mycotoxins, the relative retention times (by HPLC-ESI-TOFMS and ITMS), the exact masses of the molecular ions (by TOFMS) and the masses of the product ions, including the hydrocarbon backbones (by ITMS) of the new compounds, strongly suggested their structures.

Fumonisin measurement from maize samples by high-performance liquid chromatography coupled with corona charged aerosol detector.

Fumonisins are a class of mycotoxins produced mainly by Fusarium species, which is primary fungal contaminant of the maize and maize-derived products around the world. The B-series fumonisins (FB1, FB2 and FB3) are the most abundant and toxic constituent; thus, their levels are regulated generally worldwide. In this study, we developed a reliable method for the measurement of fumonisin FB1, FB2 and FB3 mycotoxins from maize samples without the time-consuming derivatization step using a high-performance liquid chromatograph coupled with corona charged aerosol detector. The detection and quantitation limit of the whole method were 0.02 and 0.04 mg/kg for each fumonisins, respectively. The detection linearity was tested in the calibration range of 2 orders of magnitude and the recoveries from the spiked samples were determined. The developed method proved to be sufficient to measure the maximum residue levels of fumonisins, which are specified in European Union and United States in maize and maize-based products.

Translocation and degradation of tebuconazole and prothioconazole in wheat following fungicide treatment at flowering.

BACKGROUND: Prothioconazole and tebuconazole are among the most effective fungicides against Fusarium head blight (FHB) of wheat (Triticum aestivum L.). The translocation between the ears and the flag leaves and the kinetics of degradation may influence field efficacy of these active ingredients (AIs). RESULTS: In greenhouse experiments, only traces (<1%) of the total AI content translocated from the flag leaves to the ears, and a maximum of 3.55% from the ears to the flag leaves. From the treated to the non-treated side of the ears, 3.2-15.9% of the AI translocated, depending on cultivar, AI and time. In field experiments, the degradation kinetics in the first 8 days after treatment revealed a higher velocity in the flag leaf blades than in the ears, although both were dependent on the type of cultivar. The fungicide treatment resulted in 42.6-100% decreases in FHB traits. CONCLUSIONS: There is no effective translocation of these AIs, only moderate redistribution in the ears, which can be decisive from the aspect of FHB management. The degradation of prothioconazole was faster than that of tebuconazole. Cultivar and environmental effects influenced the degradation kinetics of these AIs, but a high level of protection against FHB was maintained.

Role of fungicides, application of nozzle types, and the resistance level of wheat varieties in the control of Fusarium head blight and deoxynivalenol.

Fungicide application is a key factor in the control of mycotoxin contamination in the harvested wheat grain. However, the practical results are often disappointing. In 2000-2004, 2006-2008 and 2007 and 2008, three experiments were made to test the efficacy of fungicide control on Fusarium Head Blight (FHB) in wheat and to find ways to improve control of the disease and toxin contamination. In a testing system we have used for 20 years, tebuconazole and tebuconazole + prothioconazole fungicides regularly reduced symptoms by about 80% with a correlating reduction in toxin contamination. Averages across the years normally show a correlation of r = 0.90 or higher. The stability differences (measured by the stability index) between the poorest and the best fungicides are about 10 or more times, differing slightly in mycotoxin accumulation, FHB index (severity) and Fusarium damaged kernels (FDK). The weak fungicides, like carbendazim, were effective only when no epidemic occurred or epidemic severity was at a very low level. Similar fungicide effects were seen on wheat cultivars which varied in FHB resistance. In this study, we found three fold differences in susceptibility to FHB between highly susceptible and moderately resistant cultivars when treated with fungicides. In the moderately resistant cultivars, about 50% of the fungicide treatments lowered the DON level below the regulatory limit. In the most susceptible cultivars, all fungicides failed to reduce mycotoxin levels low enough for grain acceptance, in spite of the fact that disease was significantly reduced. The results correlated well with the results of the large-scale field tests of fungicide application at the time of natural infection. The Turbo FloodJet nozzle reduced FHB incidence and DON contamination when compared to the TeeJet XR nozzle. Overall, the data suggest that significant decreases in FHB incidence and deoxynivalenol contamination in field situations are possible with proper fungicide applications. Additionally, small plot tests can be used to evaluate the quality of the field disease and toxin production.

Detection and characterization of twenty-eight isomers of fumonisin B1 (FB1) mycotoxin in a solid rice culture infected with Fusarium verticillioides by reversed-phase high-performance liquid chromatography/electrospray ionization time-of-flight and ion trap mass spectrometry.

Fumonisin mycotoxins which are hazardous to humans and animals were produced in a Fusarium verticillioides-infected solid rice culture. To decrease the possibility of the formation of artifacts, the fumonisins were analysed by reversed-phase high-performance liquid chromatography/electrospray ionization time-of-flight (RP-HPLC/ESI-TOFMS) and ion trap mass spectrometry (RP-HPLC/ESI-ITMS) immediately after the extraction of the culture material, without any further sample clean-up. The fumonisin isomers were separated by using a flat gradient on a special, high-coverage C(18), narrow-bore HPLC column (YMC-Pack J'sphere ODS H80) suggested for the separation of structural isomers by the manufacturer. Exact mass measurements (TOFMS) of the protonated molecules and extraction of the ion chromatogram corresponding to the empirical formula (C(34)H(59)NO(15)) of FB(1) toxins led to the identification of 29 peaks and shoulders, including those of FB(1). The FB(1) toxin and 28 of its isomers were also detected by ITMS after separation with RP-HPLC. The characteristic m/z values of the product ions, including the backbones obtained by ITMS(2), undoubtedly indicated the structures of the FB(1) isomers for 28 peaks and shoulders. In the MS(2) spectra of the protonated molecules of the FB(1) isomers, with some exceptions, 15 characteristic product ions including the hydrocarbon backbone at m/z 299 were observed. The abundance ratio of the cation at m/z 299 ranged up to 5.8%. The relative quantities of the isomers found in the sample extract were expressed as percentages of the FB(1) content (0.001-0.579%). The total amount of the 28 FB(1) isomers was 2.803% of the quantity of FB(1) that is important from the aspect of food and feed safety.

Detection of new fumonisin mycotoxins and fumonisin-like compounds by reversed-phase high-performance liquid chromatography/electrospray ionization ion trap mass spectrometry.

Fumonisins were produced in a rice culture infected with Fusarium verticillioides. To decrease the possibility of the formation of artifacts, the fumonisins were analyzed by reversed-phase high-performance liquid chromatography with electrospray ionization ion trap tandem mass spectrometry (RP-HPLC/ESI-IT-MS2) immediately after the extraction of the culture material without any sample clean-up. In addition to already known fumonisins, numerous new fumonisin mycotoxins and fumonisin-like compounds were detected. On the basis of the IT-MS2 data, detailed fragmentation pathways including new mechanisms were proposed for the different series of fumonisins. The retention times, the masses of the protonated molecules and of the product ions including the backbones and the characteristic neutral mass losses from the protonated molecules of the new compounds suggested their structures (applying the well-known designation): iso-FA1a,b, iso-FB1a-d, iso-FB2,3a-e, PHFB2a-c, PHFB4a-d, FB5/iso-FB5a-d, FBK1 2TCA, FBK4 2TCA, FC2, iso-FC2,3, PHFC4, FD and FBX series. The relative quantities of fumonisins and fumonisin-like compounds found in the sample extract were expressed as percentages of FB1 (0.02-100%). The backbone of the compound denoted FD contained fewer carbon atoms than the well-known fumonisins with the C19 or C20 backbone and may well be a precursor of the longer compounds. For the compounds denoted FBX (12 compounds), one or two OH groups attached to the fumonisin backbone were esterified by carboxylic acids other than tricarballylic acid, such as cis-aconitic acid, oxalylsuccinic acid and oxalylfumaric acid.