Anne E. Desjardins

Deoxynivalenol-nonproducing Fusarium graminearum Causes Initial Infection, but does not Cause DiseaseSpread in Wheat Spikes

Fusarium graminearum is a major pathogen that causes fusarium head blight (FHB) in wheat and produces deoxynivalenol (DON) in infected grain. In previous studies, the trichodiene synthase gene (Tri5) in the fungal strain GZ3639 was disrupted to produce the DON-nonproducing strain GZT40.In this report, the virulence of strains GZ3639 and GZT40 was tested on wheat cultivars with various resistance levels by using methods of spray inoculation and injection inoculation with fungal conidia. Under field and greenhouse conditions, strain GZ3639 produced significantly more disease symptoms and reduced more yield than strain GZT40 in all wheat cultivars tested. Conidia of strain GZT40 germinated and infected inoculated spikelets, but disease symptoms were limited to inoculated spikelets without spread to uninoculated spikelets. When strain GZT40 was inoculated using the spray method, multiple initial infection sites in a spike resulted in higher levels of disease symptoms than in spikes inoculated by a single injection. Greenhouse tests confirmed that strain GZT40 did not produce DON in the infected kernels following either inoculation method. The results confirm that DON production plays a significant role in the spread of FHB within a spike, and are the first report that DON production is not necessary for initial infection by the fungus.

Distribution of Fumonisins in Maize Ears Infected with Strains of Fusarium moniliforme that Differ in Fumonisin Production

Strains of Fusarium moniliforme (Gibberella fujikuroi mating population A) that differ in fu-monisin production in vitro were previously identified in a Kansas field population. One strain that produced high levels of fumonisins and two strains that produced very low levels of fu-monisins were applied to maize kernels at planting at the Rocky Ford Farm near Manhattan, Kansas. The distribution of fumonisins in symptomatic and symptomless kernels from individual harvested ears was determined by high performance liquid chromatography, and the distribution of the three applied strains in the kernels was determined by vegetative compatibility group analysis. Both symptomatic and symptomless kernels were extensively colonized with F. moniliforme, but the highest levels of fumonisins were in the symptomatic kernels. All three applied strains were recovered from kernels in 1993, and two of them were recovered from kernels in 1994. However, a high frequency of ear and kernel infection with a strain that produced little fumonisin in vitro did not consistently decrease the level of fumonisins. The frequency of infection with fumonisin low-producing strains may have been too low for competitive exclusion of naturally occurring fumonisin high-producing strains. Also, strains that are low-fumonisin producers under laboratory conditions may be high producers in the field.

Cryptic speciation in Fusarium subglutinans

Fusarium isolates that form part of the Gibberella fujikuroi species complex have been classified using either a morphological, biological, or phylogenetic species concept. Problems with the taxonomy of Fusarium species in this complex are mostly experienced when the morphological and biological species concepts are applied. The most consistent identifications are obtained with the phylogenetic species concept. Results from recent studies have presented an example of discordance between the biological and phylogenetic species concepts, where a group of F. subglutinans sensu stricto isolates, i.e., isolates belonging to mating population E of the G. fujikuroi complex, could be sub-divided into more than one phylogenetic lineage. The aim of this study was to determine whether this sub-division represented species divergence or intraspecific diversity in F. subglutinans. For this purpose, we included 29 F. subglutinans isolates belonging to the E-mating population that were collected from either maize or teosinte, from a wide geographic range. DNA sequence data for six nuclear regions in each of these isolates were obtained and used in phylogenetic concordance analyses. These analyses revealed the presence of two major groups representing cryptic species in F. subglutinans. These cryptic species were further sub-divided into a number of smaller groups that appear to be reproductively isolated in nature. This suggests not only that the existing F. subglutinans populations are in the process of divergence, but also that each of the resulting lineages are undergoing separation into distinct taxa. These divergences did not appear to be linked to geographic origin, host, or phenotypic characters such as morphology.

Inhibition of trichothecene toxin biosynthesis by naturally occurring shikimate aromatics

Certain naturally occurring flavonoids and furanocoumarins are inhibitors of trichothecene toxin biosynthesis. These compounds block T-2 biosynthesis in liquid cultures of Fusarium sporotrichioides NRRL 3299 at concentrations substantially less than required to block fungal growth. Inhibited cultures accumulate variable amounts of trichodiene, the hydrocarbon precursor of the trichothecenes. These inhibitors appear to block the trichothecene biosynthetic pathway after formation of trichodiene and before formation of highly oxygenated trichothecenes. Exposure to these widely occurring plant shikimate aromatics may inhibit trichothecene production during plant pathogenesis.

Gibberella fujikuroi mating population E is associated with maize and teosinte.

Summary Isolates of Fusarium subglutinans mating population E are usually found on maize. This fungus forms part of the so-called Gibberella fujikuroi species complex. Previously, F. subglutinans has been associated with two additional mating populations (B and H) and a variety of plant hosts. This was mainly due to a lack of diagnostic morphological characters, but the use of DNA sequence information showed that the strains making up mating populations B, E and H, as well as those associated with the different plant hosts, represent separate species. Recently, another putative mating population has been reported on the wild teosinte relatives of maize. Based on sexual compatibility studies, these isolates were apparently closely related to the pitch canker fungus, F. subglutinans f. sp. pini (= F. circinatum;G. fujikuroi mating population H). The aim of the current study was to determine whether the population of F. subglutinans from teosinte constitutes a new or an existing lineage within the G. fujikuroi complex. For this purpose, portions of the mitochondrial small subunit ribosomal DNA, calmodulin and beta-tubulin genes from the fungi were sequenced. Phylogenetic analyses and comparison with sequences from public domain databases indicated that the F. subglutinans isolates from teosinte are most closely related to strains of G. fujikuroi mating population E. These results were confirmed using sexual compatibility studies. The putative mating population from the wild relatives of maize therefore forms part of the existing E-mating population and does not constitute a new lineage in the G. fujikuroi species complex.

Tolerance and metabolism of furanocoumarins by the phytopathogenic fungus Gibberella pulicaris (Fusarium sambucinum)

Abstract Sixty-two strains of Gibberella pulicaris (anamorph: Fusarium sambucinum) from diseased plants and from soil were tested for tolerance of the furanocoumarin xanthotoxin in vitro. Twenty-one (88%) of the plant-derived strains and two (5%) of the soil-derived strains were highly tolerant of xanthotoxin. Sixteen selected strains were tested further against 16 furanocoumarins or furanocoumarin precursors. All plant-derived strains tested were highly tolerant of and, in most cases, able to completely metabolize all 16 compounds. Most soil-derived strains tested were tolerant of furanocoumarin precursors but sensitive to certain furanocoumarins. Linear compounds methoxylated at C-8 appeared more toxic than both those unsubstituted and those with longer-chain ethers. Tolerance of angelicin, xanthotoxin, pimpinellin and isopimpinellin correlated in large part with their metabolism. All strains that were highly virulent on Pastinaca sativa root were tolerant of xanthotoxin, which is corroboration that xanthotoxin is a phytoalexin in P. sativa.

Gibberella ear rot of maize (Zea mays) in Nepal: Distribution of the mycotoxins nivalenol and deoxynivalenol in naturally and experimentally infected maize

The fungus Fusarium graminearum (sexual stage Gibberella zeae) causes ear rot of maize (Zea mays) and contamination with the 8-ketotrichothecenes nivalenol (1) or 4-deoxynivalenol (2), depending on diversity of the fungal population for the 4-oxygenase gene (TRI13). To determine the importance of 1 and 2 in maize ear rot, a survey of naturally contaminated maize in Nepal was combined with experiments in the field and in a plant growth room. In the survey, 1 contamination was 4-fold more frequent than 2 contamination and 1-producers (TRI13) were isolated more than twice as frequently as 2-producers (Psi TRI13). In maize ear rot experiments, genetically diverse 1-producers and 2-producers caused ear rot and trichothecene contamination. Among strains with the same genetic background, however, 1-producers caused less ear rot and trichothecene contamination than did 2-producers. The high frequency of 1 contamination and the high virulence of many 1-producers are of concern because maize is a staple food of rural populations in Nepal and because 1 has proven to be more toxic than 2 to animals.

Trichothecene biosynthesis inGibberella pulicaris: Inheritance of C−8 hydroxylation

SummaryNaturally occurring strains ofGibberella pulicaris (Fusarium sambucinum) produce different kinds and levels of trichothecene toxins. Progeny from crosses between strains which produce trichothecenes with an oxygen-containing group at C−8 (C8+) and those that do not (C8−) can segregate in a 1∶1 ratio for this trait. These results define a genetic locus, which we have designatedTox1. The segregation patterns observed for progeny obtained from crosses between high-toxin producers and low-toxin producers indicate that the level of toxin production is determined by several loci. One gene which controls quantitative aspects of toxin production segregates independently of both theTox1 locus and another locus which controls toxin levels. These results suggest that multiple, unlinked nuclear loci are involved in the control of trichothecene biosynthesis.

Biotransformation of the potato phytoalexin, lubimin, by Gibberella pulicaris. Identification of major products

Abstract Gibberella pulicaris (anamorph: Fusarium sambucinum) strain R-7715, biotransformed a potato phytoalexin, lubimin, into several compounds. Products forming between 1 and 6 h after addition of lubimin to cultures were two isomers of isolubimin, isomeric 15-dihydrolubimin, cyclodehydroisolubimin, and two isomers of the previously undescribed 2-dehydrolubimin. After 1–2 days, the novel tricyclic compounds, cyclolubimin (2-dihydrocyclodehydroisolubimin) and 11,12-epoxycyclo-dehydroisolubimin, became an increasing proportion of the mixture at the expense of the early products. Incubation of lubimin with the fungus in the presence of 2H2O resulted in labeling at carbon-3 of cyclodehydroisolubimin, indicating that cyclization was directed toward the double bond of a hypethetical precursor, 3,4-dehydroisolubimin, an early product tentatively identified by its mass spectrum.

5-(2-carboxyethyl)-6-hydroxy-7-methoxybenzofuran, a fungal metabolite of xanthotoxin

Tolerant strains of Gibberella pulicaris (anamorph: Fusarium sambucinum) metabolize the phytoalexin xanthotoxin to 5-(2-carboxyethyl)-6-hydroxy-7-methoxybenzofuran. This compound does not inhibit the growth of strains that are sensitive to xanthotoxin itself and all xanthotoxin-tolerant and xanthotoxin-sensitive strains tested were able to further degrade the metabolite. This is the first report of fungal detoxification of a furanocoumarin.