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Fusarium head blight (FHB) is a major disease of wheat and barley worldwide and is caused by different species in the genus Fusarium, Fusarium graminearum being the most important. We conducted population genomics analyses using SNPs obtained through genotyping by sequencing of over 500 isolates of F. graminearum from the US Upper Midwest, New York, Louisiana, and Uruguay. PCA and STRUCTURE analyses group our isolates into four previously described populations: NA1, NA2, Southern Louisiana (SLA) and Gulf Coast (GC). Some isolates were not assigned to populations because of mixed ancestry. Population structure was associated with toxin genotype and geographic origin. The NA1, NA2, and SLA populations are differentiated (F ST 0.385 - 0.551) but the presence of admixed isolates indicates that the populations are not reproductively isolated. Patterns of linkage disequilibrium (LD) decay suggest frequent recombination within populations. Fusarium graminearum populations from the US have great evolutionary potential given the high recombination rate and a large proportion of admixed isolates. The NA1, NA2, and Southern Louisiana (SLA) populations separated from their common ancestral population roughly at the same time in the past and are evolving with moderate levels of subsequent gene flow between them. Genome-wide selection scans in all three populations revealed outlier regions with the strongest signatures of recent positive natural selection. These outlier regions include many genes with unknown function and some genes with known roles in plant-microbe interaction, fungicide/drug resistance, cellular transport and genes that are related to cellular organelles. Only a very small proportion of outlier regions are shared as outliers among the three populations, suggesting unique host-pathogen interactions and environmental adaptation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Postharvest management is critical to attaining household food, nutrition, and income security. Hermetic grain storage bags offer an effective pesticide-free way to protect stored grain against fungal and insect infestation. We evaluated articles indexed in the Web of Science that included experiments comparing the storage efficacy of conventional and hermetic storage bags based on grain germination rate, insect infestation, physical damage, mycotoxin contamination, and changes in weight and moisture content. Compared with grain stored in hermetic bags, grain stored in conventional bags lost 3.6-fold more seed viability, contained 42-fold more insects, had 11-fold more physical damage, and lost 23-fold more grain weight, while grain moisture levels were similar for both hermetic and conventional storage bags. Mycotoxin contamination levels were not as frequently assessed. Levels could be low in grain stored in both types of bags, or levels could be low in hermetic bags and significantly higher in conventional bags. The improved properties of grain stored in hermetic bags can increase food security and household income by providing safe storage options for maintaining seed germinability, and for consumption and/or sale when food supplies are high, or when prices are low. Hermetic bags are economically feasible for use by subsistence farmers in Sub-Saharan Africa for grain for household consumption and for carrying-over seed for planting in the next season. Additional studies are needed to verify the mycotoxin contamination results and to determine if there are differences in functional food characteristics, e.g. flavor and cooking properties, that have not been as comprehensively studied. © 2023 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry., (© 2023 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.)

Foodborne mycotoxins are a significant food safety risk in developing countries. Our objective was to determine the occurrence of and exposure levels to aflatoxins (AFs) and fumonisins (FBs) in maize intended for human and animal consumption in food-insecure regions of western Honduras. Total AFs and FBs were quantified with a monoclonal antibody-based affinity spectrofluorimetric method. FBs were detected in 614/631 samples of maize destined for human consumption at 0.3 to 41 mg/kg (mean, 2.7 mg/kg). Of the 614 positive samples, 147 had FB levels exceeding the U.S. Food and Drug Administration (FDA) advisory threshold of 4.0 mg/kg. AFs were detected in 109/631 samples of maize for human consumption with concentrations between 1.0 and 490 µg/kg (mean, 10 µg/kg). AF levels in 34 samples exceeded the FDA regulatory limit (i.e., 20 µg/kg). The average probable daily intake of AFs in western Honduras ranged from 0 to 260 ng/kg body weight/day, and for FBs, the average probable daily intake ranged from 17 to 53 μg/kg body weight/day. AFs and FBs co-occurred in 106/631 samples with 60 samples containing both toxins at levels greater than the FDA regulatory levels. Samples of maize intended for animal feed had significantly higher AF (mean, 22 µg/kg) and FB (mean, 7.6 mg/kg) contamination levels than those observed in samples destined for human consumption. Thus, the maize supply chain in western Honduras is contaminated with mycotoxins at levels that pose health risks to both humans and livestock. More effective mycotoxin surveillance and implementation of effective mitigation strategies are needed to reduce mycotoxin contamination and exposure.

Nearly 700,000 tonnes of peanuts are consumed annually in Europe. In the last 5 years, peanuts imported from China exceeded legal European Union (EU) aflatoxin limits more than 180 times. To prevent and mitigate aflatoxin contamination, the stages of the peanut chain most vulnerable to contamination must be assessed to determine how to interrupt the movement of contaminated produce. This paper discusses effective approaches for early identification and proactive mitigation of aflatoxins in peanuts to reduce a contaminant that is an impediment to trade. We consider (i) the results of the EU Commission's Directorate-General (DG) for Health and Food Safety review, (ii) the Code of Practice for the prevention and reduction of aflatoxins in peanuts issued by Food and Agriculture Organization/World Health Organization, (iii) the results from previous EU-China efforts, and (iv) the latest state-of-the-art technology in pre- and postharvest methods as essential elements of a sustainable program for integrated disease and aflatoxin management. These include preharvest use of biocontrol, biofertilizers, improved tillage, forecasting, and risk monitoring based on analysis of big data obtained by remote sensing. At the postharvest level, we consider rapid testing methods along the supply chain, Decision Support Systems for effective silo management, and effective risk monitoring during drying, storage, and transport. Available guidance and current recommendations are provided for successful practical implementation. Food safety standards also influence stakeholder and consumer trust and confidence, so we also consider the results of multiactor stakeholder group discussions., (© 2022 The Authors. Comprehensive Reviews in Food Science and Food Safety published by Wiley Periodicals LLC on behalf of Institute of Food Technologists.)

Many species in the Fusarium fujikuroi Species Complex (FFSC) have an affinity for grass species, with whom they live in an endophytic association or cause disease. We recovered isolates of Fusarium from agriculturally important grasses in Africa and Brazil, and characterized them with morphological markers, mating type, and Amplified Fragment Length Polymorphisms (AFLPs). We also conducted multi-locus phylogenetic analyses based on partial DNA sequences of translation elongation factor-1α (TEF1), β-tubulin (TUB), and the second largest subunit of RNA polymerase (RPB2) gene regions. Sexual cross fertility was used to test the biological species concept and the sexual stage of F. madaense is described. A novel species within the FFSC, Fusarium mirum, that is different from the other known species in the complex, was formally described. Fusarium mirum, F. madaense, and Fusarium andiyazi are a tightly intertwined species trio that are morphologically identical, but phylogenetically distinguishable, and amongst whom interspecific genetic exchange may still occur. These three species are so close that they cannot be reliably distinguished if only sequences of the TEF1 gene are used. In pathogenicity tests, all tested isolates of F. madaense from sugarcane, sorghum, maize, millet and Brachiaria could induce stalk rot in sorghum, maize and millet, and pokkah boeng in sugarcane. This study increases our understanding of the diversity of species within the FFSC that cause disease in tropical grasses or act as endophytes, and their geographic distributions. The genetically close relationship between F. mirum, F. madaense, and F. andiyazi provides an opportunity to study and identify factors underlying their limited inter-specific cross-fertility and sympatric speciation., Competing Interests: Declaration of competing interest None., (Copyright © 2021 British Mycological Society. All rights reserved.)

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.

Scientific communication is facilitated by a data-driven, scientifically sound taxonomy that considers the end-user's needs and established successful practice. In 2013, the Fusarium community voiced near unanimous support for a concept of Fusarium that represented a clade comprising all agriculturally and clinically important Fusarium species, including the F. solani species complex (FSSC). Subsequently, this concept was challenged in 2015 by one research group who proposed dividing the genus Fusarium into seven genera, including the FSSC described as members of the genus Neocosmospora , with subsequent justification in 2018 based on claims that the 2013 concept of Fusarium is polyphyletic. Here, we test this claim and provide a phylogeny based on exonic nucleotide sequences of 19 orthologous protein-coding genes that strongly support the monophyly of Fusarium including the FSSC. We reassert the practical and scientific argument in support of a genus Fusarium that includes the FSSC and several other basal lineages, consistent with the longstanding use of this name among plant pathologists, medical mycologists, quarantine officials, regulatory agencies, students, and researchers with a stake in its taxonomy. In recognition of this monophyly, 40 species described as genus Neocosmospora were recombined in genus Fusarium , and nine others were renamed Fusarium. Here the global Fusarium community voices strong support for the inclusion of the FSSC in Fusarium , as it remains the best scientific, nomenclatural, and practical taxonomic option available.

Fusarium subglutinans and F. temperatum are two important fungal pathogens of maize whose distinctness as separate species has been difficult to assess. We isolated strains of these species from commercial and native maize varieties in Argentina and sequenced >28,000 loci to estimate genetic variation in the sample. Our objectives were to measure genetic divergence between the species, infer demographic parameters related to their split, and describe the population structure of the sample. When analyzed together, over 30% of each species' polymorphic sites (>2,500 sites) segregate as polymorphisms in the other. Demographic modeling confirmed the species split predated maize domestication, but subsequent between-species gene flow has occurred, with gene flow from F. subglutinans into F. temperatum greater than gene flow in the reverse direction. In F. subglutinans , little evidence exists for substructure or recent selective sweeps, but there is evidence for limited sexual reproduction. In F. temperatum , there is clear evidence for population substructure and signals of abundant recent selective sweeps, with sexual reproduction probably less common than in F. subglutinans . Both genetic variation and the relative number of polymorphisms shared between species increase near the telomeres of all 12 chromosomes, where genes related to plant-pathogen interactions often are located. Our results suggest that species boundaries between closely related Fusarium species can be semipermeable and merit further study. Such semipermeability could facilitate unanticipated genetic exchange between species and enable quicker permanent responses to changes in the agro-ecosystem, e.g., pathogen-resistant host varieties, new chemical and biological control agents, and agronomic practices.

This article is to alert medical mycologists and infectious disease specialists of recent name changes of medically important species of the filamentous mold Fusarium Fusarium species can cause localized and life-threating infections in humans. Of the 70 Fusarium species that have been reported to cause infections, close to one-third are members of the Fusarium solani species complex (FSSC), and they collectively account for approximately two-thirds of all reported Fusarium infections. Many of these species were recently given scientific names for the first time by a research group in the Netherlands, but they were misplaced in the genus Neocosmospora In this paper, we present genetic arguments that strongly support inclusion of the FSSC in Fusarium There are potentially serious consequences associated with using the name Neocosmospora for Fusarium species because clinicians need to be aware that fusaria are broadly resistant to the spectrum of antifungals that are currently available.

Fusarium subglutinans and Fusarium temperatum are common maize pathogens that produce mycotoxins and cause plant disease. The ability of these species to produce beauvericin and fumonisin mycotoxins is not settled, as reports of toxin production are not concordant. Our objective was to clarify this situation by determining both the chemotypes and genotypes for strains from both species. We analyzed 25 strains from Argentina, 13 F. subglutinans and 12 F. temperatum strains, for toxin production by ultraperformance liquid chromatography mass spectrometry (UPLC-MS). We used new genome sequences from two strains of F. subglutinans and one strain of F. temperatum , plus genomes of other Fusarium species, to determine the presence of functional gene clusters for the synthesis of these toxins. None of the strains examined from either species produced fumonisins. These strains also lack Fum biosynthetic genes but retain homologs of some genes that flank the Fum cluster in Fusarium verticillioides None of the F. subglutinans strains we examined produced beauvericin although 9 of 12 F. temperatum strains did. A complete beauvericin ( Bea ) gene cluster was present in all three new genome sequences. The Bea1 gene was presumably functional in F. temperatum but was not functional in F. subglutinans due to a large insertion and multiple mutations that resulted in premature stop codons. The accumulation of only a few mutations expected to disrupt Bea1 suggests that the process of its inactivation is relatively recent. Thus, none of the strains of F. subglutinans or F. temperatum we examined produce fumonisins, and the strains of F. subglutinans examined also cannot produce beauvericin. Variation in the ability of strains of F. temperatum to produce beauvericin requires further study and could reflect the recent shared ancestry of these two species. IMPORTANCE Fusarium subglutinans and F. temperatum are sister species and maize pathogens commonly isolated worldwide that can produce several mycotoxins and cause seedling disease, stalk rot, and ear rot. The ability of these species to produce beauvericin and fumonisin mycotoxins is not settled, as reports of toxin production are not concordant at the species level. Our results are consistent with previous reports that strains of F. subglutinans produce neither fumonisins nor beauvericin. The status of toxin production by F. temperatum needs further work. Our strains of F. temperatum did not produce fumonisins, while some strains produced beauvericin and others did not. These results enable more accurate risk assessments of potential mycotoxin contamination if strains of these species are present. The nature of the genetic inactivation of BEA1 is consistent with its relatively recent occurrence and the close phylogenetic relationship of the two sister species., (Copyright © 2020 Fumero et al.)

Fusarium proliferatum causes diverse diseases of many economically important plants. The fungus produces several mycotoxins of which the fumonisins are the most toxic. Currently, deletion of key genes for mycotoxin biosynthesis is a laborious and time-consuming procedure. We developed a novel CRISPR/Cas9-based genome-editing tool for the direct delivery of preassembled Cas9 ribonucleoproteins into protoplasts of F. proliferatum. Our CRISPR-Cas9 system couples a site-specific double-strand DNA break mediated by two Cas9 ribonucleoproteins with microhomology recombination requiring only 50-bp regions flanking the target gene. This system reduces the risk of off-target mutations and minimizes the risk of altering any gene adjacent to the target region. We used this tool to delete a polyketide synthase gene (FUM1) required for fumonisin biosynthesis. The mutants generated are no longer able to produce fumonisins, confirming the key role of FUM1 in fumonisin biosynthesis. Our CRISPR-Cas9 system is an important new tool for genetic studies of Fusarium.

Sorghum is the fifth most important cereal worldwide, spreading from Africa throughout the world. It is particularly important in the semi-arid tropics due to its drought tolerance, and when cultivated in Southeast Asia commonly occurs as a second crop during the dry season. We recovered Fusarium from sorghum in Thailand and found F. proliferatum , F. thapsinum and F. verticillioides most frequently, and intermittent isolates of F. sacchari and F. beomiforme . The relatively high frequencies of F. proliferatum and F. verticillioides , suggest mycotoxin contamination, particularly fumonisins and moniliformin, should be evaluated. Genetic variation within the three commonly recovered species was characterized with vegetative compatibility, mating type, Amplified Fragment Length Polymorphisms (AFLPs), and female fertility. Effective population number ( N e ) was highest for F. verticillioides and lowest for F. thapsinum with values based on mating type allele frequencies higher than those based on female fertility. Based on AFLP genetic variation, the F. thapsinum populations were the most closely related, the F. verticillioides populations were the most distantly related, and the F. proliferatum populations were in an intermediate position. The genetic variation observed could result if F. thapsinum is introduced primarily with seed, while F. proliferatum and F. verticillioides could arrive with seed or be carried over from previous crops, e.g. , rice or maize, which sorghum is following. Confirmation of species transmission patterns is needed to understand the agricultural systems in which sorghum is grown in Southeast Asia, which are quite different from the systems found in Africa, Australia, India and the Americas.

Maize (Zea mays), sorghum (Sorghum bicolor) and pearl millet (Pennisetum glaucum) are basic staple foods for many rural or poorer communities. These crops are susceptible to plant diseases caused by multiple species of Fusarium, some of which also produce mycotoxins, including fumonisins and moniliformin that are detrimental to both humans and domesticated animals. Eighteen potentially toxigenic Fusarium strains were isolated from maize (n = 10), sorghum (n = 7) and pearl millet (n = 1) growing in the same field in Nigeria. The 17 strains from maize and sorghum were all F. proliferatum and the one strain from pearl millet was F. pseudonygamai. Under conducive conditions, the 17 F. proliferatum strains produced fumonisins, 11 in relatively large quantities (700-17,000 mg total fumonisins, i.e., FB 1  + FB 2  + FB 3 /kg culture material), and six at <45 mg/kg. Ten F. proliferatum strains produced >100 mg of moniliformin per kg culture material with a maximum of 8900 mg/kg culture material. All strains could use all grains for growth and toxin production, regardless of the host from which they were isolated. Isolates varied in the amount of toxin produced on each substrate, with toxin production a property of the strain and not the host from which the strain was recovered. However, the extent to which a toxin-producing phenotype could be altered by the grain on which the fungus was grown is consistent with subtle genetic × environment interactions that require a larger data set than the one presented here to rigorously identify. In conclusion, there is significant variation in the ability of strains of F. proliferatum to produce fumonisins and moniliformin on maize, sorghum and millet. If the amount of toxin produced on the various grains in this study reflects real-world settings, e.g., poor storage, then the consumers of these contaminated grains could be exposed to mycotoxin levels that greatly exceed the tolerable daily intakes., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Mycotoxins are major food contaminants affecting global food security, especially in low and middle-income countries. The European Union (EU) funded project, MycoKey, focuses on &ldquo;Integrated and innovative key actions for mycotoxin management in the food and feed chains&rdquo; and the right to safe food through mycotoxin management strategies and regulation, which are fundamental to minimizing the unequal access to safe and sufficient food worldwide. As part of the MycoKey project, a Mycotoxin Charter (charter.mycokey.eu) was launched to share the need for global harmonization of mycotoxin legislation and policies and to minimize human and animal exposure worldwide, with particular attention to less developed countries that lack effective legislation. This document is in response to a demand that has built through previous European Framework Projects&mdash;MycoGlobe and MycoRed&mdash;in the previous decade to control and reduce mycotoxin contamination worldwide. All suppliers, participants and beneficiaries of the food supply chain, for example, farmers, consumers, stakeholders, researchers, members of civil society and government and so forth, are invited to sign this charter and to support this initiative.

MycoKey, an EU-funded Horizon 2020 project, includes a series of "Roundtable Discussions" to gather information on trending research areas in the field of mycotoxicology. This paper includes summaries of the Roundtable Discussions on Chemical Detection and Monitoring of mycotoxins and on the role of genetics and biodiversity in mycotoxin production. Discussions were managed by using the nominal group discussion technique, which generates numerous ideas and provides a ranking for those identified as the most important. Four questions were posed for each research area, as well as two questions that were common to both discussions. Test kits, usually antibody based, were one major focus of the discussions at the Chemical Detection and Monitoring roundtable because of their many favorable features, e.g., cost, speed and ease of use. The second area of focus for this roundtable was multi-mycotoxin detection protocols and the challenges still to be met to enable these protocols to become methods of choice for regulated mycotoxins. For the genetic and biodiversity group, both the depth and the breadth of trending research areas were notable. For some areas, e.g., microbiome studies, the suggested research questions were primarily of a descriptive nature. In other areas, multiple experimental approaches, e.g., transcriptomics, proteomics, RNAi and gene deletions, are needed to understand the regulation of toxin production and mechanisms underlying successful biological controls. Answers to the research questions will provide starting points for developing acceptable prevention and remediation processes. Forging a partnership between scientists and appropriately-placed communications experts was recognized by both groups as an essential step to communicating risks, while retaining overall confidence in the safety of the food supply and the integrity of the food production chain., Competing Interests: The authors declare no conflict of interest. The funding sponsors had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript or in the decision to publish the results.

Members of the fungal genus Fusarium can produce numerous secondary metabolites, including the nonribosomal mycotoxins beauvericin (BEA) and enniatins (ENNs). Both mycotoxins are synthesized by the multifunctional enzyme enniatin synthetase (ESYN1) that contains both peptide synthetase and S-adenosyl-l-methionine-dependent N -methyltransferase activities. Several Fusarium species can produce ENNs, BEA or both, but the mechanism(s) enabling these differential metabolic profiles is unknown. In this study, we analyzed the primary structure of ESYN1 by sequencing esyn1 transcripts from different Fusarium species. We measured ENNs and BEA production by ultra-performance liquid chromatography coupled with photodiode array and Acquity QDa mass detector (UPLC-PDA-QDa) analyses. We predicted protein structures, compared the predictions by multivariate analysis methods and found a striking correlation between BEA/ENN-producing profiles and ESYN1 three-dimensional structures. Structural differences in the β strand's Asn789-Ala793 and His797-Asp802 portions of the amino acid adenylation domain can be used to distinguish BEA/ENN-producing Fusarium isolates from those that produce only ENN.

Comparisons of draft genome sequences of three geographically distinct isolates of Fusarium fujikuroi with two recently published genome sequences from the same species suggest diverse profiles of secondary metabolite production within F. fujikuroi. Species- and lineage-specific genes, many of which appear to exhibit expression profiles that are consistent with roles in host-pathogen interactions and adaptation to environmental changes, are concentrated in subtelomeric regions. These genomic compartments also exhibit distinct gene densities and compositional characteristics with respect to other genomic partitions, and likely play a role in the generation of molecular diversity. Our data provide additional evidence that gene duplication, divergence, and differential loss play important roles in F. fujikuroi genome evolution and suggest that hundreds of lineage-specific genes might have been acquired through horizontal gene transfer., (© The Author(s) 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

Wheat is at peak quality soon after harvest. Subsequently, diverse biota use wheat as a resource in storage, including insects and mycotoxin-producing fungi. Transportation networks for stored grain are crucial to food security and provide a model system for an analysis of the population structure, evolution, and dispersal of biota in networks. We evaluated the structure of rail networks for grain transport in the United States and Eastern Australia to identify the shortest paths for the anthropogenic dispersal of pests and mycotoxins, as well as the major sources, sinks, and bridges for movement. We found important differences in the risk profile in these two countries and identified priority control points for sampling, detection, and management. An understanding of these key locations and roles within the network is a new type of basic research result in postharvest science and will provide insights for the integrated pest management of high-risk subpopulations, such as pesticide-resistant insect pests.

Aflatoxins are highly toxic carcinogens that contaminate crops worldwide. Previous studies conducted in Nigeria and Ghana found high concentrations of aflatoxins in pre- and post-harvest maize. However, little information is available on the population structure of Aspergillus Sect. Flavi in West Africa. We determined the incidence of Aspergillus Sect. Flavi and the level of aflatoxin contamination in 91 maize samples from farms and markets in Nigeria and Ghana. Aspergillus spp. were recovered from 61/91 maize samples and aflatoxins B1 and/or B2 occurred in 36/91 samples. Three samples from the farms also contained aflatoxin G1 and/or G2. Farm samples were more highly contaminated than were samples from the market, in terms of both the percentage of the samples contaminated and the level of mycotoxin contamination. One-hundred-and-thirty-five strains representative of the 1163 strains collected were identified by using a multilocus sequence analysis of portions of the genes encoding calmodulin, β-tubulin and actin, and evaluated for aflatoxin production. Of the 135 strains, there were 110 - Aspergillus flavus, 20 - Aspergillus tamarii, 2 - Aspergillus wentii, 2 - Aspergillus flavofurcatus, and 1 - Aspergillus parvisclerotigenus. Twenty-five of the A. flavus strains and the A. parvisclerotigenus strain were the only strains that produced aflatoxins. The higher contamination of the farm than the market samples suggests that the aflatoxin exposure of rural farmers is even higher than previously estimated based on reported contamination of market samples. The relative infrequency of the A. flavus SBG strains, producing small sclerotia and high levels of both aflatoxins (B and G), suggests that long-term chronic exposure to this mycotoxin are a much higher health risk in West Africa than is the acute toxicity due to very highly contaminated maize in east Africa., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Fusarium tupiense, the main causal agent of mango malformation in Brazil, is described through a combination of morphological, biological and molecular markers. This new species belongs to the Gibberella fujikuroi species complex (GFSC) and has an anamorph morphologically similar to Fusarium mangiferae and F. sterilihyphosum. F. tupiense can be differentiated from other species in the G. fujikuroi species complex on the basis of sexual crosses, amplified fragment length polymorphism (AFLP) markers and partial sequences of the tef1 and tub2 genes. Female fertility for field isolates of F. tupiense appears to be low. PCR with primers specific for the mating type (MAT) alleles and sexual crosses identified this species as heterothallic with two idiomorphs. Female-fertile tester strains were developed for the identification of field strains of this species through sexual crosses.

Acacia koa (koa), a native tree in Hawaii, suffers from a dieback caused by Fusarium oxysporum. Pathogenicity tests, vegetative compatibility group (VCG) tests, and amplified fragment length polymorphisms (AFLP) analyses were conducted on Fusarium isolates recovered from diseased koa. Koa seedling mortality with individual strains ranged from 0 to 85%, with 42% of the strains killing no seedlings. Thus, strains of F. oxysporum recovered from dying koa trees may or may not be virulent. In addition to F. oxysporum, F. pseudocircinatum strains were isolated from diseased koa; however, they were either nonvirulent or had weak virulence. This is the first report of F. pseudocircinatum in Hawaii. The 46 strains of F. oxysporum and F. pseudocircinatum were grouped into 16 VCGs, but 86% of the highly virulent strains belonged to VCG 2. In AFLP analyses, strains from the same VCG generally clustered with one another. Identification of the same set of strains using VCG, AFLP, and pathogenicity tests showed that the highly virulent strains are genetically close and that high virulence toward koa is not a property of all strains of F. oxysporum. Thus, VCG 2 with the corresponding AFLP data is a significant biological entity for which we propose the name F. oxysporum f. sp. acaciae to reflect its virulence on koa.

Fusarium graminearum (Gibberella zeae) is an important pathogen of wheat, maize, barley, and rice in South Korea, and harvested grain often is contaminated with trichothecenes such as deoxynivalenol and nivalenol. In this study, we examined 568 isolates of F. graminearum collected from maize at eight locations in South Korea. We used amplified fragment length polymorphisms (AFLPs) to identify four lineages (2, 3, 6, and 7); lineage 7 was the most common (75%), followed by lineage 6 (12%), lineage 3 (12%), and lineage 2 (1%). The genetic identity among populations was high (>0.98), and the effective migration rate between locations was higher than that between lineages. Female fertility varied by lineage: all lineage 7 isolates were fertile, while 70%, 26%, and 14% of the isolates in lineages 6, 3, and 2, respectively, were fertile. All lineage 3 and lineage 7 isolates produced deoxynivalenol, whereas most lineage 2 and 6 isolates produced nivalenol. Genotypic diversity in lineage 3 and lineage 6 populations is similar to that found in previously described Korean rice populations, but genotypic diversity in lineage 7 is much lower, even though similar levels of gene flow occur between lineage 7 populations. We conclude that lineage 7 was relatively recently introduced into South Korea, perhaps accompanying imported maize seeds.

Finger millet (Eleusine coracana) is a subsistence crop grown in Sub-Saharan Africa and the Indian Sub-continent. Fusarium species occurring on this crop have not been reported. Approximately 13% of the Fusarium isolates recovered from finger millet growing at three different locations in eastern Uganda belong to Fusarium verticillioides, and could produce up to 18,600 µg/g of total fumonisins when cultured under laboratory conditions. These strains are all genetically unique, based on AFLP analyses, and form fertile perithecia when crossed with the standard mating type tester strains for this species. All but one of the strains is female-fertile and mating-type segregates 13:20 Mat-1:Mat-2. Three new sequences of the gene encoding translation elongation factor 1-α were found within the population. These results indicate a potential health risk for infants who consume finger millet gruel as a weaning food, and are consistent with the hypothesis that F. verticillioides originated in Africa and not in the Americas, despite its widespread association with maize grown almost anywhere worldwide.

Gibberella zeae (anamorph: Fusarium graminearum) is the most common cause of Fusarium head blight (FHB) of wheat (Triticum aestivum) worldwide. Aggressiveness is the most important fungal trait affecting disease severity and stability of host resistance. Objectives were to analyze in two field experiments (i) segregation for aggressiveness among 120 progenies from each of two crosses of highly aggressive parents and (ii) stability of FHB resistance of seven moderately to highly resistant winter wheat cultivars against isolates varying for aggressiveness. Aggressiveness was measured as FHB severity per plot, Fusarium exoantigen absorbance, and deoxynivalenol content. In the first experiment, mean FHB ratings were 20 to 49% across environments and progeny. Significant genotypic variation was detected in both crosses (P < 0.01). Isolate-environment interaction explained approximately half of the total variance. Two transgressive segregants were found in cross B across environments. Traits were significantly (P < 0.05) intercorrelated. In the second experiment, despite significant (P < 0.05) genotypic variance for cultivar and isolate, no significant (P > 0.05) interaction was observed for any trait. In conclusion, progeny of highly aggressive parents might exhibit increased aggressiveness due to recombination and may, therefore, adapt nonspecifically to increased quantitative host resistance.

Fusarium graminearum is an important fungal pathogen of cereal crops and produces mycotoxins, such as the trichothecenes nivalenol and deoxynivalenol. This species may be subdivided into a series of genetic lineages or phylogenetic species. We identified strains of F. graminearum from the Republic of Korea to lineage, tested their ability to produce nivalenol and deoxynivalenol, and determined the genetic composition and structure of the populations from which they were recovered. Based on amplified fragment length polymorphism (AFLP), PCR genotyping, and chemical analyses of trichothecenes, all 249 isolates from southern provinces belonged to lineage 6, with 241 having the nivalenol genotype and 8 having the deoxynivalenol genotype. In the eastern Korea province, we recovered 84 lineage 6 isolates with the nivalenol genotype and 23 lineage 7 isolates with the deoxynivalenol genotype. Among 333 lineage 6 isolates, 36% of the AFLP bands were polymorphic, and there were 270 multilocus haplotypes. Genetic identity among populations was high (>0.972), and genotype diversity was low (30 to 58%). To test the adaptation of lineage 6 to rice, conidial mixtures of strains from lineages 3, 6, and 7 were inoculated onto rice plants and then recovered from the rice grains produced. Strains representing lineages 6 and 7 were recovered from inoculated spikelets at similar frequencies that were much higher than those for the strain representing lineage 3. Abundant perithecia were produced on rice straw, and 247 single-ascospore isolates were recovered from 247 perithecia. Perithecia representing lineage 6 (87%) were the most common, followed by those representing lineage 7 (13%), with perithecia representing lineage 3 not detected. These results suggest that F. graminearum lineage 6 may have a host preference for rice and that it may be more fit in a rice agroecosystem than are the other lineages present in Korea.

In heterothallic ascomycete fungi, idiomorphic alleles at the MAT locus control two sex pheromone-receptor pairs that function in the recognition and chemoattraction of strains with opposite mating types. In the ascomycete Gibberella zeae, the MAT locus is rearranged such that both alleles are adjacent on the same chromosome. Strains of G. zeae are self-fertile but can outcross facultatively. Our objective was to determine if pheromones retain a role in sexual reproduction in this homothallic fungus. Putative pheromone precursor genes (ppg1 and ppg2) and their corresponding pheromone receptor genes (pre2 and pre1) were identified in the genomic sequence of G. zeae by sequence similarity and microsynteny with other ascomycetes. ppg1, a homolog of the Saccharomyces alpha-factor pheromone precursor gene, was expressed in germinating conidia and mature ascospores. Expression of ppg2, a homolog of the a-factor pheromone precursor gene, was not detected in any cells. pre2 was expressed in all cells, but pre1 was expressed weakly and only in mature ascospores. ppg1 or pre2 deletion mutations reduced fertility in self-fertilization tests by approximately 50%. Deltappg1 reduced male fertility and Deltapre2 reduced female fertility in outcrossing tests. In contrast, Deltappg2 and Deltapre1 had no discernible effects on sexual function. Deltappg1/Deltappg2 and Deltapre1/Deltapre2 double mutants had the same phenotype as the Deltappg1 and Deltapre2 single mutants. Thus, one of the putative pheromone-receptor pairs (ppg1/pre2) enhances, but is not essential for, selfing and outcrossing in G. zeae whereas no functional role was found for the other pair (ppg2/pre1).

We previously published a genetic map of Gibberella zeae (Fusarium graminearum sensu lato) based on a cross between Kansas strain Z-3639 (lineage 7) and Japanese strain R-5470 (lineage 6). In this study, that genetic map was aligned with the third assembly of the genomic sequence of G. zeae strain PH-1 (lineage 7) using seven structural genes and 108 sequenced amplified fragment length polymorphism markers. Several linkage groups were combined based on the alignments, the nine original linkage groups were reduced to six groups, and the total size of the genetic map was reduced from 1,286 to 1,140 centimorgans. Nine supercontigs, comprising 99.2% of the genomic sequence assembly, were anchored to the genetic map. Eight markers (four markers from each parent) were not found in the genome assembly, and four of these markers were closely linked, suggesting that >150 kb of DNA sequence is missing from the PH-1 genome assembly. The alignments of the linkage groups and supercontigs yielded four independent sets, which is consistent with the four chromosomes reported for this fungus. Two proposed heterozygous inversions were confirmed by the alignments; otherwise, the colinearity of the genetic and physical maps was high. Two of four regions with segregation distortion were explained by the two selectable markers employed in making the cross. The average recombination rates for each chromosome were similar to those previously reported for G. zeae. Despite an inferred history of genetic isolation of lineage 6 and lineage 7, the chromosomes of these lineages remain homologous and are capable of recombination along their entire lengths, even within the inversions. This genetic map can now be used in conjunction with the physical sequence to study phenotypes (e.g., fertility and fitness) and genetic features (e.g., centromeres and recombination frequency) that do not have a known molecular signature in the genome.

Crown rot, caused by the fungus Fusarium pseudograminearum (teleomorph Gibberella coronicola) is a major disease of wheat in the Australian grain belt. However, there is little information available on the population structure of this pathogen. We measured genetic diversity as assessed with amplified fragment length polymorphism (AFLP) analysis within and between populations of F. pseudograminearum from northeastern, south central, and southwestern regions of the Australian grain belt. Amongst the 217 isolates, 176 haplotypes were identified and grouped into two main clusters. One cluster contained isolates from populations in northeastern Australia, and the other cluster contained isolates from populations in south central and southwestern Australia. The southern populations were distinguished from the northeastern populations by higher levels of population differentiation (Gst) between them and genetic identity amongst the regional populations. We hypothesize that the F. pseudograminearum populations from northeastern and southern Australia are independent, which could result from different founding events or from geographic isolation and the accumulation of genetic differences due to genetic drift and/or selection.

Genetic variation occurs at all levels across the genus Fusarium. In some cases such variation has been used to define species, and in others to describe populations or lineages. When amplified fragment length polymorphisms (AFLPs) are evaluated, strains in different species usually share at least 60% of the fragments and those in different species 40% of the fragments, or less, with isolates sharing between 40 and 60% of the fragments in an indeterminant situation. This gray area also is reflected in morphological characters, usually indistinguishable, and cross-fertility, usually some cross-fertility but often not as fertile as are strains that are more closely related. In terms of DNA sequence, the genes used for species diagnostics often have not been tested on large numbers of strains. For example, the TRI101 gene of F. graminearum contains at least 25 single nucleotide polymorphisms (SNPs) from 36 strains and yielded 17 alleles that have been proposed as a means to subdivide this species into at least nine. However these subdivisions fare poorly as more strains are analyzed, with the number of alleles increasing to >40 when approximately 500 strains from Korea and South America are sequenced. Some of the newly identified alleles cannot be correctly assigned to one of the nine subdivisions based on the proposed diagnostic SNPs. Before SNPs are proposed as characters to define species, it is important to verify their specificity based on a sufficiently large sample and to evaluate the genetic variation present in terms of an independent measure of genetic relationships. Only in such a manner can names that are meaningful in the context of trade and quarantine regulations be developed.

Mycotoxin producing capability greatly varies within species. In theory, the major source of this variability is meiotic recombination. However, a number of important toxigenic species have no known sexual stage and, therefore, the origin of the intraspecific diversity in these fungi is poorly understood. Mating in sexually reproducing Ascomycetes is controlled by MAT genes, but fungi with no known sexual stage also may have fully functional, constitutively transcribed mating type genes. The MAT genes, MAT1-1-1 and MAT1-2-1 encode putative transcription factors which, besides regulating pheromone and pheromone receptor genes, may affect other genes not involved directly in the mating process. By comparing the transcript profiles of a DeltaMAT1-2-1 knock-out mutant and the wild type of Fusarium verticillioides, more than 200 ESTs, either down- or up-regulated in the mutant, were identified. Sequences encoding proteins involved in protein synthesis and metabolism occurred more frequently among ESTs up-regulated in the mutant, while sequences involved in cell signaling and communication were more frequent in the down-regulated subset of ESTs. The lack of fertility in fungi with no known sexual stage and the limited fertility of local populations of sexually reproducing fungi are probably due to changes in one or more of numerous genes that cause female sterility. A number of gene disruption mutants of Fusarium proliferatum were assessed for their mating capabilities. Fphch (a Het-C homologue), Fpmtr (an amino acid transporter gene), and Fpnitr1 (a putative nitrilase encoding gene) encode proteins in seemingly unrelated pathways, but mutations at any of these loci can reduce female fertility. Thus, a number of genes, with functions not related directly to mating, can influence the frequency of sexual reproduction indicating that this process requires the concerted operation of many factors not obviously connected to female fertility/sterility.

Aflatoxins are a common contaminant of cereals that can cause cancer, liver disease, immune suppression, retarded growth and development, and death, depending on the level and duration of exposure. Maize is an introduced crop to Africa and there have been efforts over the last 20 years or so to replace traditional cereal crops, such as sorghum (Sorghum bicolor) and pearl millet (Pennisetum glaucum), with maize. We found that maize was significantly more heavily colonized by aflatoxin-producing Aspergillus spp. than either sorghum or millet, with overall aflatoxin levels being correspondingly higher. On average, Nigerians consume 138 kg cereals annually. If the primary cereal is sorghum instead of maize, then the risk of aflatoxin-related problems is reduced 4-fold; if it is pearl millet, then the risks are reduced 8-fold. Development programs and other ventures to increase maize production in marginal cropping areas of Africa should be reconsidered and, instead, efforts to improve/maintain traditional crops encouraged.

Fpmtr1, an amino acid transporter gene from Fusarium proliferatum was strongly expressed during conidial germination and repressed in late stationary phase. To identify the specific function of this gene, DeltaFpmtr1 knock-out mutants were generated by gene replacement. Vegetative growth of the DeltaFpmtr1 mutants was normal both in liquid and on solid media, but conidial germination was delayed. The DeltaFpmtr1 mutants and the wild type were equally fertile when used as males in sexual crosses, however if the mutants were used as the female parent then the fertility of the cross decreased dramatically. Inactivation of Fpmtr1 abolished vegetative self-incompatibility in strain ITEM 2287 of F. proliferatum, but the DeltaFpmtr1 mutants were still vegetatively incompatible with the other strains of the fungus. Endophytic colonization capability of the mutants, assessed on maize seedlings also was adversely affected. These data suggest that Fpmtr1 is involved in multiple developmental processes related to both sexual and parasexual events in F. proliferatum. Furthermore, the fungus might have problems in adapting to a less than optimal environment if this otherwise dispensable transporter has been inactivated.

For two fungal strains to be vegetatively compatible and capable of forming a stable vegetative heterokaryon they must carry matching alleles at a series of loci variously termed het or vic genes. Cloned het/vic genes from Neurospora crassa and Podospora anserina have no obvious functional similarity and have various cellular functions. Our objective was to identify the homologue of the Neurospora het-c gene in Fusarium proliferatum and to determine if this gene has a vegetative compatibility function in this economically important and widely dispersed fungal pathogen. In F. proliferatum and five other closely related Fusarium species we found a few differences in the DNA sequence, but the changes were silent and did not alter the amino acid sequence of the resulting protein. Deleting the gene altered sexual fertility as the female parent, but it did not alter male fertility or existing vegetative compatibility interactions. Replacement of the allele-specific portion of the coding sequence with the sequence of an alternate allele in N. crassa did not result in a vegetative incompatibility response in transformed strains of F. proliferatum. Thus, the fphch gene in Fusarium appears unlikely to have the vegetative compatibility function associated with its homologue in N. crassa. These results suggest that the vegetative compatibility phenotype may result from convergent evolution. Thus, the genes involved in this process may need to be identified at the species level or at the level of a group of species and could prove to be attractive targets for the development of antifungal agents.

ABSTRACT Gibberella zeae, causal agent of Fusarium head blight (FHB) of wheat and barley and Gibberella ear rot (GER) of corn, may be transported over long distances in the atmosphere. Epidemics of FHB and GER may be initiated by regional atmospheric sources of inoculum of G. zeae; however, little is known about the origin of inoculum for these epidemics. We tested the hypothesis that atmospheric populations of G. zeae are genetically diverse by determining the genetic structure of New York atmospheric populations (NYAPs) of G. zeae, and comparing them with populations of G. zeae collected from seven different states in the northern United States. Viable, airborne spores of G. zeae were collected in rotational (lacking any apparent within-field inoculum sources of G. zeae) wheat and corn fields in Aurora, NY in May through August over 3 years (2002 to 2004). We evaluated 23 amplified fragment length polymorphism (AFLP) loci in 780 isolates of G. zeae. Normalized genotypic diversity was high (ranging from 0.91 to 1.0) in NYAPs of G. zeae, and nearly all of the isolates in each of the populations represented unique AFLP haplotypes. Pairwise calculations of Nei's unbiased genetic identity were uniformly high (>0.99) for all of the possible NYAP comparisons. Although the NYAPs were genotypically diverse, they were genetically similar and potentially part of a large, interbreeding population of G. zeae in North America. Estimates of the fixation index (G(ST)) and the effective migration rate (Nm) for the NYAPs indicated significant genetic exchange among populations. Relatively low levels of linkage disequilibrium in the NYAPs suggest that outcrossing is common and that the populations are not a result of a recent bottleneck or invasion. When NYAPs were compared with those collected across the United States, the observed genetic identities between the populations ranged from 0.92 to 0.99. However, there was a significant negative correlation (R = -0.59, P < 0.001) between genetic identity and geographic distance, suggesting that some genetic isolation may occur on a continental scale. The contribution of long-distance transport of G. zeae to regional epidemics of FHB and GER remains unclear, but the diverse atmospheric populations of G. zeae suggest that inoculum may originate from multiple locations over large geographic distances. Practically, the long-distance transport of G. zeae suggests that management of inoculum sources on a local scale, unless performed over extensive production areas, will not be completely effective for the management of FHB and GER.

Fumonisin mycotoxins are commonly found on maize and pose a health risk to humans and domesticated animals. Visible sorting of grain has been suggested as a simple technique that can be used to reduce exposure to fumonisins. We collected maize samples in 2003 from different farms in the Kaduna state of Nigeria (Northern Guinea Savanna agroecological zone) that had been sorted by farmers as either good quality or poor quality. The amount of fumonisins and the presence of Fusarium verticillioides were determined for each sample. All 13 poor quality samples and the 5 good quality samples positive for fumonisins contained F. verticillioides. Twelve of 13 poor quality samples contained fumonisins (1.4 to 110 microg/g), as did the five good quality samples that were positive for F. verticillioides (0.2 to 3.7 microg of fumonisins per g). Thus, the visible sorting of grain as a technique to reduce the exposure of subsistence farmers to fumonisins could be successful if there were enough good quality grain available to permit the poor quality grain to be used for another purpose or discarded.

ABSTRACT Fusarium species are a significant component of the set of fungi associated with cassava root rot. Yield losses due to root rot average 0.5 to 1 ton/ha but losses >3 ton/ha, an equivalent of 15 to 20% yield, often occur. This paper reviews previous work on cassava root rot and summarizes a few recent studies on Fusarium species associated with the disease. Our studies in Cameroon showed that 30% of rotted tubers were infected by Fusarium spp. 12 months after planting and represented 25% of all the fungal isolates recovered. Other commonly recovered fungi were Botryodiplodia theobromae and Armillaria spp. Numerous and diverse species of Fusarium were associated with rotted cassava roots in Nigeria and Cameroon. At least 13 distinct amplified fragment length polymorphism (AFLP) groups of Fusarium were distinguishable, each group probably a distinct species, and many of them might represent previously undescribed Fusarium species. The two largest of the AFLP groups correspond to F. oxysporum and F. solani species complex. The distribution of Fusarium spp. varied among countries and among locations within a country, suggesting that germ plasm resistant at one location may not be resistant at another. Fusarium spp. also cause seedling blight of cassava and can be recovered from the stems of infected plants up to 1 m above the ground. Therefore, the pathogen can spread with stems cut as planting material. Fusarium spp. also can colonize Chromolaena odorata, the dominant weed in short fallows, which could further complicate management efforts by serving as an alternative host for strains that colonize cassava.

Six strains of Fusarium verticillioides, two of F. oxysporum, one strain of F. proliferatum, and a strain of an unidentified species were cultured on maize patties and rice and evaluated for their ability to simultaneously produce fumonisin B (FB) and C (FC) series analogues. Fumonisins were quantified by LC-MS-MS using positive ion electrospray ionization. FC1 provided characteristic fragment ions at m/z 690, 672, 654, 532, 514, and 338 corresponding to sequential loss of H2O and tricarboxylic acid moieties from the alkyl backbone, while FC3 and FC4 provided equivalent product ions 16 and 32 amu lower than the corresponding FC1 fragments, respectively. All isolates cultured on maize produced FC4. All isolates except for that of F. proliferatum also produced FC1, and three of the six strains of F. verticillioides produced FC3. All isolates except those of F. oxysporum produced detectable amounts of FB1, FB2, and FB3. Isolates that produced fumonisin B analogues produced at least 10 fold more of the B series analogues than they did of the C series analogues. The results confirm that at least some strains of F. oxysporum produce FC, but not FB, fumonisin analogues and also suggest that the genetics and physiological regulation of fumonisin production may be more complicated than previously envisaged since some strains of F. verticillioides and F. proliferatum as well as the strain of the unidentified species can simultaneously produce both FB and FC analogues.

We described the teleomorph of Fusarium sacchari as Gibberella sacchari, sp. nov. This species can be separated from other species of Gibberella on the basis of the longer, narrower ascospores found in G. sacchari and by sexual cross fertility. Female-fertile mating type tester strains were developed that can be used for making sexual crosses with this heterothallic fungus under laboratory conditions. The anamorph, Fusarium sacchari, was neotypified.

ABSTRACT Fusarium isolates recovered from sorghum and millet are commonly identified as F. moniliforme, but with the recognition of new species in this group, the strains given this name are being re-evaluated. We analyzed five strains each from five Fusarium species (F. andiyazi, F. nygamai, F. pseudonygamai, F. thapsinum, and F. verticillioides) often associated with sorghum and millet for their ability to produce fumonisin and moniliformin, their toxicity to ducklings, and their ability to cause disease on sorghum seedlings in vitro. These species can be distinguished with isozymes (fumarase, NADP-dependent isocitrate dehydrogenase, and malate dehydrogenase) and with banding patterns resulting from amplified fragment length polymorphisms. Two species, F. pseudonygamai and F. thapsinum, produced high levels of moniliformin, but little or no fumonisins, and were consistently highly toxigenic in the duckling tests. Two species, F. verticillioides and F. nygamai, produced high levels of fumonisins, but little or no moniliformin, and also were toxigenic in the duckling tests. F. andiyazi produced little or no toxin and was the least toxigenic in the duckling tests. In sorghum seedling pathogenicity tests, F. thapsinum was the most virulent followed by F. andiyazi, then F. verticillioides, and finally F. nygamai and F. pseudonygamai, which were similar to each other. Thus, these five species, which would once have all been called F. moniliforme, differ sufficiently in terms of plant pathogenicity and toxin production profile, that their previous misidentification could explain inconsistencies in the literature and differences observed by researchers who thought they were all working with the same fungal species.

Cephalosporium maydis is an important plant pathogen whose phylogenetic position relative to other fungi has not been established clearly. We compared strains of C. maydis, strains from several other plant-pathogenic Cephalosporium spp. and several possible relatives within the Gaeumannomyces-Harpophora species complex, to which C. maydis has been suggested to belong based on previous preliminary DNA sequence analyses. DNA sequences of the nuclear genes encoding the rDNA ITS region, β-tubulin, histone H3, and MAT-2 support the hypothesis that C. maydis is a distinct taxon within the Gaeumannomyces-Harpophora species complex. Based on amplified fragment length polymorphism (AFLP) profiles, C. maydis also is distinct from the other tested species of Cephalosporium, Phialophora sensu lato and members of Gaeumannomyces-Harpophora species complex, which supports its classification as Harpophora maydis. Oligonucleotide primers for H. maydis were developed that can be used in a PCR diagnostic protocol to rapidly and reliably detect and identify this pathogen. These diagnostic PCR primers will aid the detection of H. maydis in diseased maize because this fungus can be difficult to detect and isolate, and the movement of authentic cultures may be limited by quarantine restrictions.

ABSTRACT Gibberella zeae is the major fungal pathogen of Fusarium head blight of wheat and produces several mycotoxins that are harmful to humans and domesticated animals. We identified loci associated with pathogenicity and aggressiveness on an amplified fragment length polymorphism 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 for 2 years. Progeny segregated qualitatively (61:38) for pathogenicity:nonpathogenicity, respectively. The trait maps to linkage group IV, which is 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 using simple interval analysis. 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 that was 50 centimorgans away explained 29% of the phenotypic variation. TRI5 is tightly linked to the locus controlling trichothecene toxin type. The two QTLs, however, were likely part of the same QTL using composite interval analysis. Progeny that produced deoxynivalenol were, on average, approximately twice as aggressive as those that produced 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.

Fusarium species from agricultural crops have been well studied with respect to toxin production and genetic diversity, while similar studies of communities from nonagricultural plants are much more limited. We examined 72 Fusarium isolates from a native North American tallgrass prairie and found that Gibberella intermedia (Fusarium proliferatum), Gibberella moniliformis (Fusarium verticillioides), and Gibberella konza (Fusarium konzum) dominated. Gibberella thapsina (Fusarium thapsinum) and Gibberella subglutinans (Fusarium subglutinans) also were recovered, as were seven isolates that could not be assigned to any previously described species on the basis of either morphological or molecular characters. In general, isolates from the prairie grasses produced the same toxins in quantities similar to those produced by isolates of the same species recovered from agricultural hosts. The G. konza isolates produce little or no fumonisins (up to 120 micro g/g by one strain), and variable but generally low to moderate amounts of beauvericin (4 to 320 micro g/g) and fusaproliferin (50 to 540 micro g/g). Toxicity to Artemia salina larvae within most species was correlated with the concentration of either beauvericin or fusaproliferin produced. Organic isolates from some cultures of G. moniliformis were highly toxic towards A. salina even though they produced little, if any, beauvericin or fusaproliferin. Thus, additional potentially toxigenic compounds may be synthesized by G. moniliformis strains isolated from prairie grasses. The Fusarium community from these grasses appears to contain some species not found in surrounding agricultural communities, including some that probably are undescribed, and could be capable of serving as a reservoir for strains of potential agricultural importance.

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. 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. We think that 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.

The Gibberella fujikuroi species complex (Fusarium section Liseola and allied taxa) is composed of an increasingly large number of morphological, biological and phylogenetic species. Most of the known species in this group have been isolated from agricultural ecosystems or have been described from a small number of isolates. We sampled Fusarium communities from native prairie grasses in Kansas and recovered a large number of isolates that superficially resemble F. anthophilum. We used a combination of morphological, biological and molecular characters to describe a new species, Gibberella konza (Gibberella fujikuroi mating population I [MP-I]), from native prairie grasses in Kansas. Although female fertility for field isolates of this species appears to be low, G. konza is heterothallic, and we developed reliably female fertile mating population tester strains for this species. The F. konzum anamorph is differentiated from F. anthophilum and from other Fusarium species in section Liseola by mating compatibility, morphology, AFLP fingerprint profile and differences in β-tubulin DNA sequence.

ABSTRACT Gibberella zeae (anamorph Fusarium graminearum) causes Fusarium head blight (FHB) of wheat and barley and has been responsible for several billion dollars of losses in the United States since the early 1990s. We isolated G. zeae from the top, middle, and bottom positions of wheat spikes collected from 0.25-m(2) quadrats during severe FHB epidemics in a single Kansas (KS) field (1993) and in a single North Dakota (ND) field (1994). Three amplified fragment length polymorphism (AFLP) primer pairs were used to resolve 94 polymorphic loci from 253 isolates. Members of a subset of 26 isolates also were tested for vegetative compatibility groups (VCGs). Both methods indicated high levels of genotypic variability and identified the same sets of isolates as probable clones. The mean number of AFLP multilocus haplotypes per head was approximately 1.8 in each population, but this value probably underestimates the true mean due to the small number of samples taken from each head. Isolates with the same AFLP haplotype often were recovered from different positions in a single head, but only rarely were such apparently clonal isolates recovered from more than one head within a quadrat, a pattern that is consistent with a genetically diverse initial inoculum and limited secondary spread. The KS and ND samples had no common AFLP haplotypes. All G. zeae isolates had high AFLP fingerprint similarity (>70%, unweighted pair group method with arithmetic means similarity) to reference isolates of G. zeae lineage 7. The genetic identity between the KS and ND populations was >99% and the estimated effective migration rate was high (Nm approximately 70). Tests for linkage disequilibrium provide little evidence for nonrandom associations between loci. Our results suggest that these populations are parts of a single, panmictic population that experiences frequent recombination. Our results also suggest that a variety of population sampling designs may be satisfactory for assessing diversity in this fungus.

ABSTRACT Cephalosporium maydis, the causal agent of late wilt of maize, was first described in Egypt in the 1960s, where it can cause yield losses of up to 40% in susceptible plantings. We characterized 866 isolates of C. maydis collected from 14 governates in Egypt, 7 in the Nile River Delta and 7 in southern (Middle and Upper) Egypt, with amplified fragment length polymorphism (AFLP) markers. The four AFLP primer-pair combinations generated 68 bands, 25 of which were polymorphic, resulting in 52 clonal haplotypes that clustered the 866 isolates into four phylogenetic lineages. Three lineages were found in both the Nile River Delta and southern Egypt. Lineage IV, the most diverse group (20 haplotypes), was recovered only from governates in the Nile River Delta. In some locations, one lineage dominated (up to 98% of the isolates recovered) and, from some fields, only a single haplotype was recovered. Under field conditions in Egypt, there is no evidence that C. maydis reproduces sexually. The nonuniform geographic distribution of the pathogen lineages within the country could be due to differences in climate or in the farming system, because host material differs in susceptibility and C. maydis lineages differ in pathogenicity.

Thirty-six Fusarium strains were grown on cracked yellow corn and evaluated for optimum fusaproliferin production, with Fusarium subglutinans E-1583 producing the highest levels (1600 microg/g). Three solvent systems were tested for extracting fusaproliferin from the cultures of F. subglutinans E-1583. Methanol gave the highest fusaproliferin recovery, followed by methanol/1% aqueous NaCl (55:45, v/v) and acetonitrile/methanol/H(2)O (16:3:1, v/v/v). Hexane partitioning was effective in removing many impurities from the crude fusaproliferin extracts prior to the liquid chromatography step. Fusaproliferin samples were further purified by high-performance liquid chromatography (HPLC) with a C18 preparatory column using a mobile phase of acetonitrile/H(2)O (80:20, v/v). The purity of the fusaproliferin was verified by analytical HPLC, GC/MS, (1)H NMR spectroscopy, and electrospray ionization (ESI) MS. The isolated fusaproliferin was shown to be free of impurities and can be used as a standard for routine analysis. Fusaproliferin was shown to be temperature-sensitive when samples were stored at room temperature (20-24 degrees C) for more than several days. After 30 days at 4 degrees C, approximately 8% of the fusaproliferin had been transformed to deacetyl-fusaproliferin; however, samples stored at -20 degrees C for 1 year contained only trace amounts of the deacetylated form.

Fifteen Fusarium species were analyzed by high-performance liquid chromatography for the production of six mycotoxins in corn grits cultures. Production of mycotoxins ranged from 66 to 2,500 micro g/kg for fumonisin B(1), 0.6 to 1,500 micro g/g for moniliformin, 2.2 to 720 micro g/g for beauvericin, and 12 to 130 micro g/g for fusaproliferin. Fumonisin B(2) (360 micro g/kg) was produced by two species, fumonisin B(3) was not detected in any of the 15 species examined, and Fusarium bulbicola produced none of the six mycotoxins that we analyzed.