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A-to-I RNA editing catalyzed by adenosine-deaminase-acting-on-RNA (ADARs) was assumed to be unique to metazoans because fungi and plants lack ADAR homologs. However, genome-wide messenger RNA (mRNA) editing was found to occur specifically during sexual reproduction in filamentous ascomycetes. Because systematic characterization of adenosine/cytosine deaminase genes has implicated the involvement of TAD2 and TAD3 orthologs in A-to-I editing, in this study, we used genetic and biochemical approaches to characterize the role of FgTAD2, an essential adenosine-deaminase-acting-on-tRNA (ADAT) gene, in mRNA editing in Fusarium graminearum. FgTAD2 had a sexual-stage-specific isoform and formed heterodimers with enzymatically inactive FgTAD3. Using a repeat-induced point (RIP) mutation approach, we identified 17 mutations in FgTAD2 that affected mRNA editing during sexual reproduction but had no effect on transfer RNA (tRNA) editing and vegetative growth. The functional importance of the H352Y and Q375*(nonsense) mutations in sexual reproduction and mRNA editing were confirmed by introducing specific point mutations into the endogenous FgTAD2 allele in the wild type. An in vitro assay was developed to show that FgTad2-His proteins purified from perithecia, but not from vegetative hyphae, had mRNA editing activities. Moreover, the H352Y mutation affected the enzymatic activity of FgTad2 to edit mRNA but had no effect on its ADAT activity. We also identified proteins co-purified with FgTad2-His by mass spectrometry analysis and found that two of them have the RNA recognition motif. Taken together, genetic and biochemical data from this study demonstrated that FgTad2, an ADAT, catalyzes A-to-I mRNA editing with the stage-specific isoform and cofactors during sexual reproduction in fungi. [ABSTRACT FROM AUTHOR]

Key points Efficient procedures for production and regeneration of Gibberella zeae protoplasts were determined. Fungal mutants were obtained with elevated 7β-hydroxylase activity. Mutant G. zeae M23 almost fully converts LCA (4 g/L) to UDCA.

The establishment of the harmful pathogen Fusarium graminearum in different agroecosystems may strongly depend on the ability of the soils to suppress its development and survival. This study aimed to evaluate the influence of different soil tillage systems (i.e., conventional tillage, reduced tillage and no-tillage) on soil fungistasis against F. graminearum. Soil samples were collected three times during the plant growing season in 2016 and 2017 from a long-term, 20-year soil tillage experiment. The F. graminearum in the soil samples was quantified by real-time qPCR. The soil fungistasis was evaluated by the reduction in the radial growth of F. graminearum in an in vitro assay. The antagonistic activity of the soil bacteria was tested using the dual culture method. The F. graminearum DNA contents in the soils were negatively correlated with soil fungistasis (r = –0.649 *). F. graminearum growth on the unfumigated soil was reduced by 70–87% compared to the chloroform fumigated soil. After the plant vegetation renewal, the soil fungistasis intensity was higher in the conventionally tilled fields than in the no-tillage. However, no significant differences were obtained among the tillage treatments at the mid-plant growth stage and after harvesting. 23 out of 104 bacteria isolated from the soil had a moderate effect, and only 1 had a strong inhibitory effect on the growth of F. graminearum. This bacterium was assigned 100% similarity to the Bacillus amyloliquefaciens Hy7 strain (gene bank no: JN382250) according to the sequence of the 16S ribosome subunit coding gene. The results of our study suggest that the presence of F. graminearum in soil is suppressed by soil fungistasis; however, the role of tillage is influenced by other factors, such as soil biological activity, type and quantity of plant residues and environmental conditions. [ABSTRACT FROM AUTHOR]

Abstract Mitogen-activated protein kinase (MAPK) cascades are activated by external stimuli and convert signals to cellular changes. Individual MAPKs have been characterized in a number of plant pathogenic fungi for their roles in pathogenesis and responses to biotic or abiotic stresses. However, mutants deleted of all the MAPK genes have not been reported in filamentous fungi. To determine the MAPK-less effects in a fungal pathogen, in this study we generated and characterized mutants deleted of all three MAPK genes in the wheat scab fungus Fusarium graminearum. The Gpmk1 mgv1 Fghog1 triple mutants had severe growth defects and was non-pathogenic. It was defective in infection cushion formation and DON production. Conidiation was reduced in the triple mutant, which often produced elongated conidia with more septa than the wild-type conidia. The triple mutant was blocked in sexual reproduction due to the loss of female fertility. Lack of any MAPKs resulted in an increased sensitivity to various abiotic stress including cell wall, osmotic, oxidative stresses, and phytoalexins, which are likely related to the defects of the triple mutant in environmental adaptation and plant infection. The triple mutant also had increased sensitivity to the biocontrol bacterium Bacillus velezensis and fungus Clonostachys rosea. In co-incubation assays with B. velezensis, the Gpmk1 mgv1 Fghog1 mutant had more severe growth limitation than the wild type and was defective in conidium germination and germ tube growth. In confrontation assays, the triple mutant was defective in defending against mycoparasitic activities of C. rosea and the latter could grow over the mutant but not wild-type F. graminearum. RNA-seq and metabolomics analyses showed that the MAPK triple mutant was altered in the expression of many ATP-binding cassette (ABC) and major facilitator superfamily (MFS) transporter genes and the accumulation of metabolites related to arachidonic acid, linoleic acid, and alpha-linolenic acid metabolisms. Overall, as the first study on mutants deleted of all three MAPKs in fungal pathogens, our results showed that although MAPKs are not essential for growth and asexual reproduction, the Gpmk1 mgv1 Fghog1 triple mutant was blocked in plant infection and sexual reproductions. It also had severe defects in responses to various abiotic stresses and bacterial- or fungal-fungal interactions.

CK1 casein kinases are well conserved in filamentous fungi. However, their functions are not well characterized in plant pathogens. In Fusarium graminearum, deletion of FgYCK1 caused severe growth defects and loss of conidiation, fertility, and pathogenicity. Interestingly, the Fgyck1 mutant was not stable and often produced fast-growing spontaneous suppressors. Suppressor mutations were frequently identified in the FgBNI4 gene by sequencing analyses. Deletion of the entire FgBNI4 or disruptions of its conserved C-terminal region could suppress the defects of Fgyck1 in hyphal growth and conidiation, indicating the genetic relationship between FgYCK1 and FgBNI4. Furthermore, the Fgyck1 mutant showed defects in polarized growth, cell wall integrity, internalization of FgRho1 and vacuole fusion, which were all partially suppressed by deletion of FgBNI4. Overall, our results indicate a stage-specific functional relationship between FgYCK1 and FgBNI4, possibly via FgRho1 signaling for regulating polarized hyphal growth and cell wall integrity. [ABSTRACT FROM AUTHOR]

Ursodeoxycholic acid (UDCA) is a highly demanded pharmaceutical steroid widely used in medicine. An ascomycete Gibberella zeae VKM F-2600 is capable of producing UDCA by 7β-hydroxylation of lithocholic acid (LCA). The present study is aimed at the improvement of the fungus productivity. The original procedures for the protoplast obtaining followed by UV mutagenesis and screening of ketoconazole-resistant mutant clones have been applied. The highest yield of G. zeae protoplasts was obtained when using the mycelium in the active growth phase, ammonium chloride as an osmotic stabilizer and treatment of the fungal cells by the lytic enzymes cocktail from Trichoderma hurzanium. The conditions for effective protoplast regeneration and the UV-mutagenesis were found to provide 6–12% survival rate of the protoplasts with superior number of possible mutations. Three of 27 ketoconazole-resistant mutant clones obtained have been selected due to their increased biocatalytic activity towards LCA. The mutant G. zeae M23 produced 26% more UDCA even at relatively high LCA concentration (4 g/L) as compared with parent fungal strain, and the conversion reached 88% (w/w). The yield of UDCA reached in this study prefers those ever reported. The results contribute to the knowledge on ascomycete mutagenesis, and are of importance for biotechnological production of value added cholic acids. Key points: Efficient procedures for production and regeneration of Gibberella zeae protoplasts were determined. Fungal mutants were obtained with elevated 7β-hydroxylase activity. Mutant G. zeae M23 almost fully converts LCA (4 g/L) to UDCA. [ABSTRACT FROM AUTHOR]

Wheat farmers rely on fungicides to protect fields against several foliar and flowering diseases, including Fusarium head blight (FHB). A range of active ingredients is used in isolation or in dual premixes that include a dimethylation inhibitor (DMI) or a quinone outside inhibitor (QoI) fungicide. Comprehensive information about fungicide resistance in F. graminearum is available for DMIs, while for QoIs the data are scarce. We characterized 225 strains obtained from two states in southern Brazil, Rio Grande do Sul (RS) and Paraná (PR), in relation to their response to two QoIs. The median EC50 (effective concentration leading to 50% inhibition of conidial germination) value for azoxystrobin (n = 25 isolates) was 2.20 μg/ml in the PR population and 4.04 μg/ml in the RS population. For pyraclostrobin (n = 50), the median EC50 was 0.28 μg/ml in the PR population and 0.24 μg/ml in the RS population. Evidence of cross‐resistance could not be detected. Screening using a discriminatory dose (DD) for azoxystrobin in a larger number of isolates from PR (n = 75) and RS (n = 100) states allowed the detection of 50% and 28% sensitive strains, respectively. Using the DD for pyraclostrobin, 33% and 18.8% were classified as less sensitive in the PR and RS isolates, respectively. In RS, the frequency of less‐sensitive isolates increased over time (2007–2011). No point mutation at any of the target spots (F129L, G137R, G143A) was detected. Our results represent an important step towards the establishment of a sensitivity profile for two of the most commonly used QoIs in commercial premixes targeting FHB control. [ABSTRACT FROM AUTHOR]

The establishment of the harmful pathogen Fusarium graminearum in different agroecosystems may strongly depend on the ability of the soils to suppress its development and survival. This study aimed to evaluate the influence of different soil tillage systems (i.e., conventional tillage, reduced tillage and no-tillage) on soil fungistasis against F. graminearum. Soil samples were collected three times during the plant growing season in 2016 and 2017 from a long-term, 20-year soil tillage experiment. The F. graminearum in the soil samples was quantified by real-time qPCR. The soil fungistasis was evaluated by the reduction in the radial growth of F. graminearum in an in vitro assay. The antagonistic activity of the soil bacteria was tested using the dual culture method. The F. graminearum DNA contents in the soils were negatively correlated with soil fungistasis (r = –0.649 *). F. graminearum growth on the unfumigated soil was reduced by 70–87% compared to the chloroform fumigated soil. After the plant vegetation renewal, the soil fungistasis intensity was higher in the conventionally tilled fields than in the no-tillage. However, no significant differences were obtained among the tillage treatments at the mid-plant growth stage and after harvesting. 23 out of 104 bacteria isolated from the soil had a moderate effect, and only 1 had a strong inhibitory effect on the growth of F. graminearum. This bacterium was assigned 100% similarity to the Bacillus amyloliquefaciens Hy7 strain (gene bank no: JN382250) according to the sequence of the 16S ribosome subunit coding gene. The results of our study suggest that the presence of F. graminearum in soil is suppressed by soil fungistasis; however, the role of tillage is influenced by other factors, such as soil biological activity, type and quantity of plant residues and environmental conditions.

Fusarium head blight (FHB), also known as head scab, is a devastating fungal disease that affects small grain cereal crops such as wheat (Triticum aestivum L.). The predominant causal agent, Fusarium graminearum Schwabe (teleomorph: Gibberella zeae (Schwein.) Petch), is ranked the fourth most important fungal plant pathogen worldwide. Apart from yield and quality losses, mycotoxin production can occur from FHB infection, resulting in harmful effects on human and animal health. Some level of disease control may be achieved by using certain fungicides and agronomic practices plus host resistance. In South Africa, there are currently no registered fungicides or bio-fungicides, no resistant wheat cultivars and only limited control is achieved by cultural practices. Because effective disease reduction cannot be achieved by using a single strategy, the integration of multiple management strategies can enhance disease control. We review possible strategies for reducing the risk for FHB infections that are relevant to the context of South Africa and other wheat growing areas in Africa. Significance: • The importance of the effect of FHB on wheat cannot be overemphasised. This review highlights and describes the various control options and their efficacies. It also describes the current state of research in an effort to control FHB and its associated mycotoxins. • Wheat is one of the most produced crops worldwide and in South Africa, hence this review could promote and intensify research towards the development of more effective management strategies for FHB of wheat.

Abstract: The objective of this work was to assess the androgenic response, via microspore culture, of wheat genotypes with different levels of resistance to Gibberella zeae. The number of androgenic embryos per spike, and of green and albino plants was counted for the BRS 179 (moderately resistant), Frontana and Sumai 3 (resistant), and BRS 194, Embrapa 27, and Fielder (susceptible) genotypes. The degree of interference by the Fielder, Pavon 76, and Sumai 3 ovary-donor genotypes, used for co-culture with the microspore cells, was also assessed regarding androgenic response. Induction efficiency ranged from 0.33 embryo per spike for Embrapa 27 to 109.8 embryos for Frontana. Sumai 3 presented the second best response, and Embrapa 27 behaved as highly recalcitrant. The co-culture of ovaries from the Fielder genotype stands out from the others and positively interferes in the embryo induction rate. No significant differences were observed for the regeneration frequency of green plants; however, for albino plants, BRS 194 produced the highest rate. Frontana and Sumai 3 present the highest androgenic response rates and can be used in breeding programs for the rapid development of cultivars resistant to Gibberella zeae.

Fusarium head blight (FHB), also known as head scab, is a devastating fungal disease that affects small grain cereal crops such as wheat (Triticum aestivum L.). The predominant causal agent, Fusarium graminearum Schwabe (teleomorph: Gibberella zeae (Schwein.) Petch), is ranked the fourth most important fungal plant pathogen worldwide. Apart from yield and quality losses, mycotoxin production can occur from FHB infection, resulting in harmful effects on human and animal health. Some level of disease control may be achieved by using certain fungicides and agronomic practices plus host resistance. In South Africa, there are currently no registered fungicides or bio-fungicides, no resistant wheat cultivars and only limited control is achieved by cultural practices. Because effective disease reduction cannot be achieved by using a single strategy, the integration of multiple management strategies can enhance disease control. We review possible strategies for reducing the risk for FHB infections that are relevant to the context of South Africa and other wheat growing areas in Africa. Significance: • The importance of the effect of FHB on wheat cannot be overemphasised. This review highlights and describes the various control options and their efficacies. It also describes the current state of research in an effort to control FHB and its associated mycotoxins. • Wheat is one of the most produced crops worldwide and in South Africa, hence this review could promote and intensify research towards the development of more effective management strategies for FHB of wheat. [ABSTRACT FROM AUTHOR]

The article presents a research report on deoxynivalenol contamination for different wheat grain handling conditions. Topics include that deoxynivalenol is a mycotoxin byproduct of Fusarium graminearum infection; and that Fusarium graminearum infection can occur during anthesis through to the soft dough stage and can continue to grow at a grain moisture content.

Fusarium head blight (FHB), caused mainly by Fusarium graminearum, is the foremost destructive disease of cereals worldwide. Effector-like molecules produced by F. graminearum play key roles in the infection process and are assumed to be one of the essential components of the pathogen’s aggressiveness. However, their nature and role in the disease are still largely misunderstood. As a mean to provide relevant information about the molecular determinism of F. graminearum aggressiveness, we surveyed three F. graminearum strains on three wheat cultivars contrasted by their susceptibility to FHB. F. graminearum strains revealed large differences in aggressiveness which were mostly unchanged when facing hosts of contrasted susceptibility, suggesting that their behavior rely on intrinsic determinants. Surveying the fungal mass progress and the mycotoxin production rate in the spikes did not evidence any simple relationship with aggressiveness differences, while clues were found through a qualitative and quantitative characterization of the three strain proteomes established in planta especially with regards to early synthesized putative effectors. Independently of the wheat cultivar, the three F. graminearum strains produced systematically the same protein set during the infection but substantial differences in their abundance enabled the categorization of fungal aggressiveness. Overall, our findings show that the contrasts in F. graminearum aggressiveness were not based on the existence of strain-specific molecules but rather on the ability of the strain to ensure their sufficient accumulation. Protein abundance variance was mostly driven by the strain genetics and part was also influenced by the host cultivar but strain by cultivar interactions were marginally detected, depicting that strain-specific protein accumulations did not depend on the host cultivar. All these data provide new knowledge on fungal aggressiveness determinants and provide a resourceful repertoire of candidate effector proteins to guide further research.

ABSTRACT: The fungus Fusarium graminearum was one of the first pathogens described as causing infections in rice; however, in Brazil, there is no description of its occurrence in panicles. The present study aimed to describe the symptoms caused by F. graminearum infection in irrigated rice grains. The experiment was conducted in a greenhouse in duplicate using the irrigated rice cultivar SCS 121CL and hybrid INOV CL at the R4 (flowering) stage. Two isolate of Fusarium graminearium species complex 15A (F. graminearium - 15-ADON) and FmNiv (F. meridionale - Nivalenol), was inoculated onto panicles by spraying with macroconidia and the development of symptoms was monitored until harvest. There was no difference in symptoms among isolates. Light brown spots were observed in the glumes three days after inoculation. These later evolved into brown lesions of irregular shape and size. The glume darkened to purple when the grains were in the filling stage (R6). On maturation, the glume showed dark brown coloration. Severely infected grains were shriveled and brittle.

A-to-I RNA editing catalyzed by adenosine-deaminase-acting-on-RNA (ADARs) was assumed to be unique to metazoans because fungi and plants lack ADAR homologs. However, genome-wide messenger RNA (mRNA) editing was found to occur specifically during sexual reproduction in filamentous ascomycetes. Because systematic characterization of adenosine/cytosine deaminase genes has implicated the involvement of TAD2 and TAD3 orthologs in A-to-I editing, in this study, we used genetic and biochemical approaches to characterize the role of FgTAD2 , an essential adenosine-deaminase-acting-on-tRNA (ADAT) gene, in mRNA editing in Fusarium graminearum . FgTAD2 had a sexual-stage-specific isoform and formed heterodimers with enzymatically inactive FgTAD3 . Using a repeat-induced point (RIP) mutation approach, we identified 17 mutations in FgTAD2 that affected mRNA editing during sexual reproduction but had no effect on transfer RNA (tRNA) editing and vegetative growth. The functional importance of the H352Y and Q375*(nonsense) mutations in sexual reproduction and mRNA editing were confirmed by introducing specific point mutations into the endogenous FgTAD2 allele in the wild type. An in vitro assay was developed to show that FgTad2-His proteins purified from perithecia, but not from vegetative hyphae, had mRNA editing activities. Moreover, the H352Y mutation affected the enzymatic activity of FgTad2 to edit mRNA but had no effect on its ADAT activity. We also identified proteins co-purified with FgTad2-His by mass spectrometry analysis and found that two of them have the RNA recognition motif. Taken together, genetic and biochemical data from this study demonstrated that FgTad2, an ADAT, catalyzes A-to-I mRNA editing with the stage-specific isoform and cofactors during sexual reproduction in fungi., Competing Interests: Competing interests statement:The authors declare no competing interest.

Fusarium head blight (FHB), caused mainly by Fusarium graminearum , is the foremost destructive disease of cereals worldwide. Effector-like molecules produced by F. graminearum play key roles in the infection process and are assumed to be one of the essential components of the pathogen's aggressiveness. However, their nature and role in the disease are still largely misunderstood. As a mean to provide relevant information about the molecular determinism of F. graminearum aggressiveness, we surveyed three F. graminearum strains on three wheat cultivars contrasted by their susceptibility to FHB. F. graminearum strains revealed large differences in aggressiveness which were mostly unchanged when facing hosts of contrasted susceptibility, suggesting that their behavior rely on intrinsic determinants. Surveying the fungal mass progress and the mycotoxin production rate in the spikes did not evidence any simple relationship with aggressiveness differences, while clues were found through a qualitative and quantitative characterization of the three strain proteomes established in planta especially with regards to early synthesized putative effectors. Independently of the wheat cultivar, the three F. graminearum strains produced systematically the same protein set during the infection but substantial differences in their abundance enabled the categorization of fungal aggressiveness. Overall, our findings show that the contrasts in F. graminearum aggressiveness were not based on the existence of strain-specific molecules but rather on the ability of the strain to ensure their sufficient accumulation. Protein abundance variance was mostly driven by the strain genetics and part was also influenced by the host cultivar but strain by cultivar interactions were marginally detected, depicting that strain-specific protein accumulations did not depend on the host cultivar. All these data provide new knowledge on fungal aggressiveness determinants and provide a resourceful repertoire of candidate effector proteins to guide further research. [ABSTRACT FROM AUTHOR]

The involvement of different parameters on Gibberella zeae lipase (GZEL) membrane binding were characterized by using monomolecular film technology and circular dichroism spectroscopy. Among four kinds of phospholipid monolayers, 1,2‑dimyristoyl‑ sn ‑glycero‑3‑phosphoethanolamine have the highest maximum insertion pressure (MIP) value. Comparing the GZEL adsorption to phosphatidylcholine monolayers with different acyl chains in sn -1 and sn -2 positions, the higher MIP values were found for 1,2‑dilauroyl‑ sn ‑glycero‑3‑phosphocholine. Significantly improvement between 1,2‑dioleoyl‑ sn ‑glycero‑3‑phosphocholine and 1,2‑distearoyl‑ sn ‑glycero‑3‑phosphocholine suggested that the presence of fatty acid unsaturation may affect protein adsorption by changing the chemical structure in each phospholipid. The MIP value was shown higher (48.6 mN m−1) at pH 5 and pH 6 (47.5 ± 1.9 mN m−1) but decreased significantly (34.2 mN m−1) at pH 9. This may indicate that the proportion of helices in the protein decreases with the alteration of the catalytic center, thus affecting the binding of the protein to its substrate. The MIP values obviously decreased with increasing salt ion concentration, suggesting that excessive salt ion concentration may destabilize the secondary and tertiary structures of the protein, thereby affecting the characteristics of its adsorption at the interfaces. Present studies improve our understanding on the protein-membrane interaction of this enzyme. • Phospholipid head group, chain length and unsaturation influence the adsorption of GZEL into phospholipid monolayers. • The pH and NaCl concentration of subphase affect the binding of GZEL to phospholipid monolayers. • Different pH can influence the secondary structure of GZEL as investigated by circular dichroism spectroscopy. [ABSTRACT FROM AUTHOR]

Fusarium graminearum and Fusarium virguliforme are two fungal pathogens that can infect soybean [Glycine max (L.) Merr.] early in the growing season. A greenhouse experiment was conducted to determine if the presence of both pathogens resulted in an increased negative impact on early soybean growth compared with each pathogen alone. Treatment factors consisted of four inoculation treatments: a non-inoculated control, inoculation with F. graminearum only (FG), inoculation with F. virguliforme only (FV), and a combination of both pathogens (FG + FV). As well, three soybean cultivars (CH2105R2, P92Y51 and 'Sloan'), and three seed treatments (a control, mefenoxam + fludioxonil, and fluxapyroxad) were evaluated. Results indicated FG + FV did not result in increased root disease, a decrease in shoot growth, or reduced root growth compared with FG and FV. The FV inoculation treatment exhibited more root disease severity compared with FG, but there were no differences between the two inoculation treatments for the other below-ground measurements, as well as all above-ground measurements. Fungicide seed treatment did not reduce root disease severity or lead to increased early soybean growth compared with the non-treated control. Cultivar differences were the primary factor which influenced early-season soybean growth. Because there was no evidence of an increased effect between presence of F. graminearum and F. virguliforme, these results suggest management decisions for both pathogens can remain separate for soybean growers in Wisconsin, but management of F. virguliforme should take priority over F. graminearum due to its prevalent cause of soybean sudden death syndrome. [ABSTRACT FROM AUTHOR]

Members of the Fusarium graminearum species complex (Fg complex or FGSC) are the primary pathogens causing Fusarium head blight in wheat and barley worldwide. A natural pathogenicity mutant (strain 0225022) was found in a sample of the Fg complex collected in Japan. The mutant strain did not induce symptoms in wheat spikes beyond the point of inoculation, and did not form perithecia. No segregation of phenotypic deficiencies occurred in the progenies of a cross between the mutant and a fully pathogenic wild-type strain, which suggested that a single genetic locus controlled both traits. The locus was mapped to chromosome 2 by using sequence-tagged markers; and a deletion of ∼3 kb was detected in the mapped region of the mutant strain. The wild-type strain contains the FGSG_02810 gene, encoding a putative glycosylphosphatidylinositol anchor protein, in this region. The contribution of FGSG_02810 to pathogenicity and perithecium formation was confirmed by complementation in the mutant strain using gene transfer, and by gene disruption in the wild-type strain.

Abstract Background The crystal structure of lipase from Gibberella zeae (GZEL) indicates that its C-terminal extension is composed of a loop and a α-helix. This structure is unique, possibly providing novel evidence on lipase mechanisms. Methods Two C-terminally truncated mutants (GZEL-Δ(α-helix) and GZEL-Δ(α-helix+loop)) were constructed. The role of these secondary structure segments on enzymatic activities and interfacial binding properties of GZEL was investigated by using conventional pH-stat method and monomolecular film techniques. In addition, inactive variants (Ser144Ala) of wild-type GZEL and two truncated mutants were constructed and produced specifically for interfacial binding experiments. Results Compared to the wild-type GZEL, lipase and phospholipase activities were significantly decreased in the two mutants. Deletion of the α-helix had great influence on the lipase activity of GZEL, resulting in residual 7.3% activity; the additional deletion of the loop led to 8.1% lipase activity. As for the phospholipase function, residual activities of 63.0% and 35.4% were maintained for GZEL-Δ(α-helix) and GZEL-Δ(α-helix+loop), respectively. Findings obtained with monomolecular film experiments further indicated that the reduction in phospholipase activity occurred with the anionic phospholipid as substrate, but was not seen with zwitterionic phospholipid. Results of the maximum insertion pressure, synergy factor and binding kinetic parameters documented that the α-helix structure of GZEL strongly influence the binding and insertion of enzyme to the phospholipid monolayer. Moreover, the interfacial binding function of α-helix was partly conformed by connecting to the C-terminal of Aspergillus oryzae lipase. General significance Our results provide important information on the understanding of the structure-function relationship of GZEL. Graphical abstract Schematic model of the interfacial binding of GZEL and its C-terminal truncated mutants (GZEL-Δ(α-helix) and GZEL-Δ(α-helix+loop)) to the membrane. Unlabelled Image Highlights • The C-terminal structure of GZEL presents unique membrane binding properties. • The deletion on the C-terminal peptides of GZEL reduces lipase and phospholipase activities. • The α-helix on C-terminal has more impact on lipase activity than phospholipase activity. • The α-helix helps GZEL absorption and further insert into the phospholipid monolayer. • The interfacial binding function of α-helix is confirmed in MdlB. [ABSTRACT FROM AUTHOR]

With the aim of unravelling the role of airborne Fusarium graminearum inoculum in the epidemic of Fusarium head blight (FHB) caused by this species in wheat spikes, a network of Burkard air samplers was set up in five wheat fields distributed in Belgium from 2011 to 2013. Each year from April to July, the daily amounts of F. graminearum inoculum above the wheat canopy were quantified using a newly developed TaqMan qPCR assay. The pattern of spore trapping observed was drastically different per year and per location with a frequency of detection between 9 and 66% and a mean daily concentration between 0.8 and 10.2 conidia-equivalent/m3. In one location, air was sampled for a whole year. Inoculum was frequently detected from the wheat stem elongation stage until the end of the harvesting period, but high inoculum levels were also observed during the fall. Using a window-pane analysis, different periods of time around wheat flowering (varying in length and starting date) were investigated for their importance in the relation between airborne inoculum and FHB parameters (FHB severity, frequency of F. graminearum infection and DON). For almost all the combinations of variables, strong and significant correlations were found for multiple window lengths and starting times. Inoculum quantities trapped around flowering were highly correlated with F. graminearum infection (up to R = 0.84) and DON (up to R = 0.9). Frequencies of detection were also well correlated with both of these parameters. DON concentrations at harvest could even be significantly associated with the F. graminearum inoculum trapped during periods finishing before the beginning of the anthesis (R = 0.77). Overall, these results highlight the key role of the airborne inoculum in F. graminearum epidemics and underline the importance of monitoring it for the development of disease forecasting tools. [ABSTRACT FROM AUTHOR]

Summary: Recombination is a major evolutionary force, increasing genetic diversity and permitting efficient coevolution of fungal pathogen(s) with their host(s). The ascomycete Fusarium graminearum is a devastating pathogen of cereal crops, and can contaminate food and feed with harmful mycotoxins. Previous studies have suggested a high adaptive potential of this pathogen, illustrated by an increase in pathogenicity and resistance to fungicides. In this study, we provide the first detailed picture of the crossover events occurring during meiosis and discuss the role of recombination in pathogen evolution. An experimental recombinant population (n = 88) was created and genotyped using 1306 polymorphic markers obtained from restriction site‐associated DNA sequencing (RAD‐seq) and aligned to the reference genome. The construction of a high‐density linkage map, anchoring 99% of the total length of the reference genome, allowed the identification of 1451 putative crossovers, positioned at a median resolution of 24 kb. The majority of crossovers (87.2%) occurred in a relatively small portion of the genome (30%). All chromosomes demonstrated recombination‐active sections, which had a near 15‐fold higher crossover rate than non‐active recombinant sections. The recombination rate showed a strong positive correlation with nucleotide diversity, and recombination‐active regions were enriched for genes with a putative role in host–pathogen interaction, as well as putative diversifying genes. Our results confirm the preliminary analysis observed in other F. graminearum strains and suggest a conserved ‘two‐speed’ recombination landscape. The consequences with regard to the evolutionary potential of this major fungal pathogen are also discussed. [ABSTRACT FROM AUTHOR]

The plant pathogen Fusarium graminearum causes Fusarium head blight in cereal crops and produces mycotoxins that are harmful to animals and humans. For the initiation and spread of disease, asexual and sexual reproduction is required. Therefore, studies on fungal reproduction contribute to the development of new methods to control and maintain the fungal population. Screening a previously generated transcription factor mutant collection, we identified one putative C₂H₂ zinc-finger transcription factor, pcs1, which is required for both sexual and asexual reproduction. Deleting pcs1 in F. graminearum resulted in a dramatic reduction in conidial production and a complete loss of sexual reproduction. The pathways and gene ontology of pcs1-dependent genes from microarray experiments showed that several G-protein related pathways, oxidase activity, ribosome biogenesis, and RNA binding and processing were highly enriched, suggesting that pcs1 is involved in several different biological processes. Further, overexpression of pcs1 increased conidial production and resulted in earlier maturation of ascospores compared to the wild-type strain. Additionally, the vegetative growth of the overexpression mutants was decreased in nutrient-rich conditions but was not different from the wild-type strain in nutrient-poor conditions. Overall, we discovered that the pcs1 transcription factor positively regulates both conidiation and sexual reproduction and confers nutrient condition-dependent vegetative growth.

The aim of the present study is to design and synthesize a series of novel 4(3 H)-quinazolinone derivatives containing an isoxazole moiety and evaluate their antifungal activity against Gibberella zeae ( G. zeae), Fusarium oxysporum ( F. oxysporum), Cytospora mandshurica ( C. mandshurica), Phytophthora infestans ( P. infestans), and Pellicularia sasakii ( P. sasakii), and their antibacterial activity against Ralstonia solanacearum ( R. solanacearum) and Xanthomomu oryzae pv. oryzae ( Xoo). Bioassay results showed that the target compounds 8a-8o had certain antifungal activities against the test fungus at the concentration of 50 μg/mL, which were lower than those of Hymexazol and Epoxiconazole. Meanwhile, preliminary bioassay results showed that compounds 8a-8o had better antibacterial activity against R. solanacearum and Xoo at 200 and 100 μg/mL. In particular, compound 8i exhibited significant antibacterial activity against Xoo in vitro, with an inhibition rate of 100% at 200 μg/mL, which was superior to those of Bismerthiazol (72%) and Thiodiazole copper (35%). [ABSTRACT FROM AUTHOR]

Using the classical emulsified system and the monomolecular film technique, the substrate specificity of recombinant Gibberella zeae lipase (rGZEL) that originates from Gibberella zeae was characterized in detail. Under the emulsified reaction system, both phospholipase and glycolipid hydrolytic activities were observed, except for the predominant lipase activity. The optimum conditions for different activity exhibition were also determined. Compared with its lipase activity, a little higher ratio of glycolipid hydrolytic activity (0.06) than phospholipase activity (0.02) was found. rGZEL preferred medium chain-length triglycerides, while lower activity was found for the longer-chain triglyceride. Using the monomolecular film technique, we found that the preference order of rGZEL to different phospholipids was 1,2-diacyl-sn-glycero-3-phospho-L-serine (PS) > 1,2-dioleoyl-sn-glycero-3-phospho-rac-(1-glycerol) sodium salt (PG) > 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) > L-α-phosphatidylinositol (PI) > cardiolipin (CL) > 3-sn-phosphatidic acid sodium salt (PA) > L-α-phosphatidylethanolamine (PE), while no hydrolytic activity was detected for sphingomyelin (SM). Moreover, rGZEL showed higher galactolipase activity on 1,2-distearoyimonoglactosylglyceride (MGDG). A kinetic study on the stereo- and regioselectivity of rGZEL was also performed by using three pairs of pseudodiglyceride enantiomers (DDGs). rGZEL presented higher preference for distal DDG enantiomers than adjacent ester groups, however, no hydrolytic activity to the sn-2 position of diglyceride analogs was found. Furthermore, rGZEL preferred the R configuration of DDG enantiomers. Molecular docking results were in concordance with in vitro tests. [ABSTRACT FROM AUTHOR]

Fusarium graminearum is a major pathogen to cause wheat scab. G protein coupled receptors (GPCRs) are a class of important cell surface receptors. The cAMP signal pathway mediated by GPCRs may be involved in pathopoiesis of F. graminearum and synthesis of mycotoxin. Studies on the structure and physiological and chemical characters of such proteins will be useful for understanding the function and effect to pathopoiesis of the protein. The cAMP receptors like GPCR genes of F. graminearum genome sequence were analyzed through bioinformatics approaches. Five typical cAMP receptors like GPCRs, FgcAR1, FgcAR2, FgcAR3, FgcAR4 and FgcAR5, were identified in F. graminearum, which contained seven trans-membrane domain structures and were located on plasma membrane. Four proteins are hydrophobin except for FgcAR1. All of them have lots of a helical with a proportion around 60%, but there is no β strand in FgcAR4 and FgcAR5 and precious few of β strand in FgcAR1, FgcAR2 and FgcAR3 with the proportion less than 3%. A certain number of phosphorylation loci of Ser and Thr are involved in the receptor. Genetic relationship analysis showed that the cAR protein in F. graiminearum has a high sequence homology and close genetic relationship with that in F. pseudograminearum and F. langsethiae. The study clarified the physiological and chemical characters, location, secondary structure, phosphorylation loci of cAMP Receptor-like GPCRs in F. graminearum and the genetic relationship in Fusarum family, which will facilitate to further understand the mechanism of pathogesis in wheat scab and develop novel fungicide by using GPCRs as drug targets. [ABSTRACT FROM AUTHOR]

A novel type of O -carboxymethyl chitosan Schiff bases ( O -CSPX) was synthesized via a condensation reaction. After the coordination reaction of cupric ions, Cu(II) complexes ( O -CSPX-Cu) were achieved. The theoretical structure of O -CSPX-Cu calculated by Gaussian 09 reveals that the copper ions underwent dsp 2 hybridization, coordinated by the carbon atom in the p-π conjugate group and the oxygen atoms in the acetate ion. Then, the structures were confirmed by FT-IR, 1 H NMR, CP-MAS 13 C NMR, elemental analysis, DSC and XRD. The antifungal properties of O -CSPX-Cu against Phytophthora capsici (P. capsici), Gibberella zeae (G. zeae) and Glomerella cingulata (G. cingulata) were evaluated at concentrations ranging from 0.05 mg/mL to 0.20 mg/mL. The experiments indicated that the derivatives have significantly enhanced antifungal activity after copper ion complexation compared with the original chitosan. Moreover, it was shown that 0.20 mg/mL of O -CSP3-Cu and O -CSP4-Cu can 100% inhibit the growth of P. capsici . The experimental results reveal that the antifungal efficiency is related to the space steric hindrance on the benzene ring, which may provide a novel direction for the development of copper fungicides. [ABSTRACT FROM AUTHOR]

The hypovirulence-inducing Fusarium graminearum virus China 9 (FgV-ch9) was described recently and is closely related to the Fusarium graminearum mycovirus-2 (FgV2). Both viruses share common properties of viruses belonging to the family Chrysoviridae. Re-sequencing of FgV-ch9 revealed duplications of the 3′ non-coding regions of segments 2 and 3. Both duplications are arranged in a head-to-tail array, are attached to the complete terminus, and do not affect the encoded gene. An internal duplication was found in segment 5. This duplication resulted in an increase in the size of the encoded protein. In silico analysis showed similar duplications in segments 2 and 3 of FgV2. [ABSTRACT FROM AUTHOR]

Fusarium graminearum is the main causal agent of Fusarium head blight (FHB) on wheat and barley. In a previous study, a GIN4-like protein kinase gene, GIL1, was found to be important for plant infection and sexual reproduction. In this study we further characterized the functions of GIL1 kinase in different developmental processes. The Dgil1 mutants were reduced in growth, conidiation, and virulence, and formed whitish and compact colonies. Although phialide formation was rarely observed in the mutants, deletion of GIL1 resulted in increased hyphal branching and increased tolerance to cell wall and cell membrane stresses. The Dgil1 mutants produced straight, elongated conidia lacking of distinct foot cells and being delayed in germination. Compared with the wild type, some compartments in the vegetative hyphae of Dgil1 mutants had longer septal distances and increased number of nuclei, suggesting GIL1 is related to cytokinesis and septation. Localization of the GIL1-GFP fusion proteins to the septum and hyphal branching and fusion sites further supported its roles in septation and branching. Overall, our results indicate that GIL1 plays a role in vegetative growth and plant infection in F. graminearum, and is involved in septation and hyphal branching. [ABSTRACT FROM AUTHOR]

No Brasil, diversas espécies de Fusarium são patogênicas ao milho, afetando sementes, plântulas, colmos e grãos. Objetivou-se comparar isolados de F. verticillioides e de duas espécies do complexo F. graminearum (F. graminearum sensu stricto e F. meridionale), de ocorrência predominante em sementes de milho, quanto à taxa de crescimento micelial, capacidade de colonização e redução da germinação da semente, redução da emergência de plântulas e infecção e colonização de colmos de milho.Testes permutacionais e análise de contraste foram usados para discriminar os isolados. Análise de componentes principais foi utilizada para identificar as variáveis responsáveis pela maior variância entre os isolados. Com exceção da redução da germinação, os isolados F. verticillioides apresentaram menores valores de taxa de crescimento micelial, eficiência de infecção, redução na emergência de plântulas e comprimento de lesão em colmos, quando comparados aos demais, que não se diferenciaram. As variáveis redução da emergência e redução da germinação apresentaram maior influência na caracterização dos isolados, sugerindo que sementes infectadas com isolados de qualquer uma das espécies do complexo F. graminearum testadas representam maior risco ao estabelecimento da cultura do que isolados de F. verticillioides.In Brazil, several Fusarium species are pathogenic to corn affecting seeds, seedlings, stalks and grains. This research aimed to compare isolates from F. verticillioides and from two species of the F. graminearum complex (F. graminearum sensu stricto and F. meridionale) prevalent on corn seeds in relation to mycelial growth rate, ability to colonize and reduce seed germination, reduction in seedling emergence and infection and colonization of corn stalk. Permutational test and contrast analysis was performed to discriminate the isolates and species. Principal component analysis was used to identify the variables leading to greatest variance among the isolates. With the exception of seed germination, F. verticillioides isolates showed lower values for mycelial growth rate, infection efficiency, reduction in seedling emergence and lesion size than F. graminearum complex isolates that were all similar. Seedling emergence and seed germination showed greater influence on the characterization of isolates, suggesting that seeds infected with F. graminearum isolates of any of the species tested represent a greater threat to crop stand than F. verticillioides isolates.

The fungus Fusarium graminearum was one of the first pathogens described as causing infections in rice; however, in Brazil, there is no description of its occurrence in panicles. The present study aimed to describe the symptoms caused by F. graminearum infection in irrigated rice grains. The experiment was conducted in a greenhouse in duplicate using the irrigated rice cultivar SCS 121CL and hybrid INOV CL at the R4 (flowering) stage. Two isolate of Fusarium graminearium species complex 15A (F. graminearium - 15-ADON) and FmNiv (F. meridionale - Nivalenol), was inoculated onto panicles by spraying with macroconidia and the development of symptoms was monitored until harvest. There was no difference in symptoms among isolates. Light brown spots were observed in the glumes three days after inoculation. These later evolved into brown lesions of irregular shape and size. The glume darkened to purple when the grains were in the filling stage (R6). On maturation, the glume showed dark brown coloration. Severely infected grains were shriveled and brittle. [ABSTRACT FROM AUTHOR]

Gibberella zeae (anamorph Fusarium graminearum) was determined on seed of St. John's wort produced in localities Pančevo, Gorobilje and Kačarevo during 2008 and 2009. Morphological characteristics of monosporial isolates were described. All tested isolates formed in vitro perithecia, and were classified as F. graminearum (syn. F. graminearum Group 2). Pathogenicity was confirmed by the artificial inoculation of St. John's wort seed.

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.

O controle químico da giberela pode ser influenciado pelo momento da aplicação e pelo fungicida. Na safra agrícola de 2004, os fungicidas foram aplicados em três estádios de crescimento da planta: início da floração, metade da floração completa e no início do desenvolvimento de grão leitoso. Em 2005, uma única aplicação foi feita no início da floração, testando-se maior número de fungicidas. As aplicações foram feitas com pulverizador manual, com pressão gerada por gás CO2, numa vazão de 200L ha-1. A intensidade da giberela foi quantificada pela incidência, severidade e índice de giberela. Também foram determinados o rendimento de grãos, o peso de mil grãos e a incidência de Fusarium graminearum nos grãos colhidos. O maior percentual de controle da giberela ocorreu com aplicação feita no início da floração. Os fungicidas metconazole (0,09L i.a. ha-1), tebuconazole (0,15), trifloxistrobina+tebuconazole (0,075+0,15), azoxistrobina+ciproconazole (0,060+0,024) e piraclostrobina+epoxiconazole (0,099+ 0,037) reduziram significativamente a intensidade da giberela, obtendo-se controle médio de 46% na incidência, 46,6% na severidade e 71,2% no índice de giberela. Esses fungicidas mostraram ganho médio no rendimento de grãos de 23,4%, porém apresentaram baixa eficácia no controle da infecção de F. graminearum nos grãos.The chemical control of the wheat scab can be affected by time of application and fungicide. In the 2004 growing season, the fungicides were applied in the plant growth stage of beginning of flowering, half of complete flowering and beginning of soft dough stage. In the 2005 growing season, one only fungicides application was done in the beginning of the flowering with a larger number of fungicides. The applications were done with manual spray, with CO2 gas pressure in an outflow of 200L ha-1. The intensity of wheat scab was quantified by the incidence, severity and index of giberela. Grain yield, weight of a thousand grains and the incidence of Fusarium graminearum in the grains were also evaluated. The high porcentual of wheat scab control occurred when the application was done in the beginning of the flowering. The fungicides metconazol (0.09L i.a. ha-1), tebuconazol (0.15), trifloxystrobin+tebuconazol (0.075+0.15), azoxystrobin+ciproconazol (0.060+0.024) and pyraclostrobin+epoxiconazol (0.099+0.037) reduced significantly the intensity of giberela wheat, obtaining average control of 46% on incidence, 46.6% on severity and 71.2% on index of giberela wheat. These fungicides showed grain yield average of 23.4%, but a low efficiency on F. graminearum infection control in grains.

Fusarium graminearum is a ubiquitous pathogen of cereal crops, including wheat, barley, and maize. Diseases caused by F. graminearum are of particular concern because harvested grains frequently are contaminated with harmful mycotoxins such as deoxynivalenol (DON). In this study, we explored the role of Ras GTPases in pathogenesis. The genome of F. graminearum contains two putative Ras GTPase-encoding genes. The two genes (RAS1 and RAS2) showed different patterns of expression under different conditions of nutrient availability and in various mutant backgrounds. RAS2 was dispensable for survival but, when disrupted, caused a variety of morphological defects, including slower growth on solid media, delayed spore germination, and significant reductions in virulence on wheat heads and maize silks. Intracellular cAMP levels were not affected by deletion of RAS2 and exogenous treatment of the ras2 mutant with cAMP did not affect phenotypic abnormalities, thus indicating that RAS2 plays a minor or no role in cAMP signaling. However, phosphorylation of the mitogen-activated protein (MAP) kinase Gpmk1 and expression of a secreted lipase (FGL1) required for infection were reduced significantly in the ras2 mutant. Based on these observations, we hypothesize that RAS2 regulates growth and virulence in F. graminearum by regulating the Gpmk1 MAP kinase pathway.