Evolution of the Toxb Gene in Pyrenophora tritici-repentis and Related Species.

Pyrenophora tritici-repentis (tan spot) is a destructive foliar pathogen of wheat with global impact. This ascomycete fungus possesses a highly plastic open pangenome shaped by the gain and loss of effector genes. This study investigated the allelic variations in the chlorosis-encoding gene ToxB across 422 isolates representing all identified pathotypes and worldwide origins. To gain better insights into ToxB evolution, we examined its presence and variability in other Pyrenophora spp. A ToxB haplotype network was constructed, revealing the evolutionary relationships of this gene (20 haplotypes) across four Pyrenophora species. Notably, toxb, the homolog of ToxB, was detected for the first time in the barley pathogen Pyrenophora teres. The ToxB/toxb genes display evidence of selection that is characterized by loss of function, duplication, and diverse mutations. Within the ToxB/toxb open reading frame, 72 mutations were identified, including 14 synonymous, 55 nonsynonymous, and 3 indel mutations. Remarkably, a, ∼5.6-kb Copia-like retrotransposon, named Copia-1_Ptr, was found inserted in the toxb gene of a race 3 isolate. This insert disrupted the ToxB gene's function, a first case of effector gene disruption by a transposable element in P. tritici-repentis. Additionally, a microsatellite with 25 nucleotide repeats (0 to 10) in the upstream region of ToxB suggested a potential mechanism influencing ToxB expression and regulation. Exploring ToxB-like protein distribution in other ascomycetes revealed the presence of ToxB-like proteins in 19 additional species, including the Leotiomycetes class for the first time. The presence/absence pattern of ToxB-like proteins defied species relatedness compared with a phylogenetic tree, suggesting a past horizontal gene transfer event during the evolution of the ToxB gene. [Formula: see text] Copyright © 2024 His Majesty the King in Right of Canada, as represented by the Minister of Agriculture and Agri-Food. This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Correction: Masi et al. Araufuranone: A New Phytotoxic Tetrasubstituted Dihydrofuro[3,2-b]furan-2(5H)-One Isolated from Ascochyta araujiae. Biomolecules 2022, 12, 1274.

In the original article [...].

SDHI resistance in Pyrenophora teres f teres and molecular detection of novel double mutations in sdh genes conferring high resistance.

BACKGROUND: Net blotch (NB), caused by Pyrenophora teres f. teres (Ptt), is an important disease of barley worldwide. NB control is commonly achieved through the use of fungicide mixtures including strobilurins, triazoles and carboxamides. Succinate dehydrogenase inhibitors (SDHI) are important components of fungicide management programs of barley diseases. However, during the last growing seasons in Argentina, barley fields sprayed with mixtures containing SDHI fungicides have shown failures in NB control. Here, we report the isolation and characterization of Argentine Ptt strains resistant to SDHI fungicides. RESULTS: Compared against a sensitive (wild-type) reference strain collected in 2008, all 21 Ptt isolates collected in 2021 exhibited resistance to pydiflumetofen and fluxapyroxad both in vitro and in vivo. Concordantly, all of them presented target-site mutations in any of the sdhB, sdhC and sdhD genes. Although the mutations detected have been previously reported in other parts of the world, this study documents for the first time the occurrence of double mutations in the same Ptt isolate. Specifically, the double mutation sdhC-N75S + sdhD-D145G confers high resistance to SDHI fungicides, while the double mutations sdhB-H277Y + sdhC-N75S and sdhB-H277Y + sdhC-H134R confer moderate levels of resistance in Ptt. CONCLUSIONS: SDHI-resistance in Argentine Ptt populations is expected to increase. These findings emphasize the urgent need to perform a wider survey and a more frequent monitoring of SDHI sensitivity of Ptt populations and to develop and implement effective antiresistance tactics. © 2023 Society of Chemical Industry.

Araufuranone: A New Phytotoxic Tetrasubstituted Dihydrofuro[3,2-b]furan-2(5H)-One Isolated from Ascochyta araujiae.

Araujia hortorum is a perennial vining plant species native to South America. It was introduced into many countries for ornamental and medicinal purposes as well as for its edible fruits, but it has become highly invasive, generating severe environmental problems. Biological control using bioherbicides and natural compounds is an interesting control option. The pathogenic fungus Ascochyta araujiae, isolated from infected leaves of A. hortorum, could be considered as a potential biocontrol agent. Its ability to produce bioactive metabolites was studied. The organic extract of the fungal culture filtrates showed interesting phytotoxic activities consisting of clearly visible necrotic symptoms (0.5-1 cm in diameter) in the punctured leaves. Thus, it was purified; this afforded three main metabolites. These were chemically and biologically characterised: one proved to be a new pentasubstituted dihydrofuro[3,2-b]furan-2(5H)-one, named araufuranone (1). The others were the already known fungal metabolites neovasinin and 2,4-dihydroxy-6-hydoxymethylbenzaldehyde (2 and 3). The structure of araufuranone was determined using spectroscopic methods (essentially 1D and 2D 1H and 13C NMR and HR ESIMS spectra); its relative configuration was assigned by a NOESY spectrum. To the best of our knowledge, araufuranone is the first example of a naturally occurring compound showing that carbon skeleton. Assayed by a puncture, araufuranone proved to be weakly active on the leaves of Diplotaxis sp. and Sonchus sp.; the other two metabolites were even less toxic. Tested on cress, compounds 2 and 3 were able to partially inhibit rootlet elongation whereas araufuranone was almost inactive.

Truncatenolide, a Bioactive Disubstituted Nonenolide Produced by Colletotrichum truncatum, the Causal Agent of Anthracnose of Soybean in Argentina: Fungal Antagonism and SAR Studies.

A bioactive disubstituted nonenolide, named truncatenolide, was produced by Colletotrichum truncatum, which was collected from infected tissues of soybean showing anthracnose symptoms in Argentina. This is a devastating disease that drastically reduces the yield of soybean production in the world. The fungus also produced a new trisubstituted oct-2-en-4-one, named truncatenone, and the well-known tyrosol and N-acetyltyramine. Truncatenolide and truncatenone were characterized by spectroscopic (essentially one-dimensional (1D) and two-dimensional (2D) 1H and 13C NMR and HR ESIMS) and chemical methods as (5E,7R,10R)-7-hydroxy-10-methyl-3,4,7,8,9,10-hexahydro-2H-oxecin-2-one and (Z)-6-hydroxy-3,5-dimethyloct-2-en-4-one, respectively. The geometry of the double bond of truncatenolide was assigned by the value of olefinic proton coupling constant and that of truncatenone by the correlation observed in the corresponding NOESY spectrum. The relative configuration of each stereogenic center was assigned with the help of 13C chemical shift and 1H-1H scalar coupling DFT calculations, while the absolute configuration assignment of truncatenolide was performed by electronic circular dichroism (ECD). When tested on soybean seeds, truncatenolide showed the strongest phytotoxic activity. Tyrosol and N-acetyltyramine also showed phytotoxicity to a lesser extent, while truncatenone weakly stimulated the growth of the seed root in comparison to the control. When assayed against Macrophomina phaseolina and Cercospora nicotianae, other severe pathogens of soybean, truncatenolide showed significant activity against M. phaseolina and total inhibition of C. nicotianae. Thus, some other fungal nonenolides and their derivatives were assayed for their antifungal activity against both fungi in comparison with truncatenolide. Pinolidoxin showed to a less extent antifungal activity against both fungi, while modiolide A selectively and totally inhibited only the growth of C. nicotianae. The SAR results and the potential of truncatenolide, modiolide A, and pinolidoxin as biofungicides were also discussed.

Pseudomonas fluorescens Showing Antifungal Activity against Macrophomina phaseolina, a Severe Pathogenic Fungus of Soybean, Produces Phenazine as the Main Active Metabolite.

Pseudomonas fluorescens 9 and Bacillus subtilis 54, proposed as biofungicides to control Macrophomina phaseolina, a dangerous pathogen of soybean and other crops, were grown in vitro to evaluate their ability to produce metabolites with antifungal activity. The aim of the manuscript was to identify the natural compounds responsible for their antifungal activity. Only the culture filtrates of P. fluorescens 9 showed strong antifungal activity against M. phaseolina. Its organic extract contained phenazine and mesaconic acid (1 and 2), whose antifungal activity was tested against M. phaseolina, as well as Cercospora nicotianae and Colletotrichum truncatum, other pathogens of soybean; however, only compound 1 exhibited activity. The antifungal activity of compound 1 was compared to phenazine-1-carboxylic acid (PCA, 3), 2-hydroxyphenazine (2-OH P, 4), and various semisynthetic phenazine nitro derivatives in order to perform a structure-activity relationship (SAR) study. PCA and phenazine exhibited the same percentage of growth inhibition in M. phaseolina and C. truncatum, whereas PCA (3) showed lower activity against C. nicotianae than phenazine. 2-Hydroxyphenazine (4) showed no antifungal activity against M. phaseolina. The results of the SAR study showed that electron attractor (COOH and NO2) or repulsor (OH) groups significantly affect the antifungal growth, as well as their α- or ß-location on the phenazine ring. Both PCA and phenazine could be proposed as biopesticides to control the soybean pathogens M. phaseolina, C. nicotianae, and C. truncatum, and these results should prompt an investigation of their large-scale production and their suitable formulation for greenhouse and field applications.

Current recommendations and novel strategies for sustainable management of soybean sudden death syndrome.

The increase in food production requires reduction of the damage caused by plant pathogens, minimizing the environmental impact of management practices. Soil-borne pathogens are among the most relevant pathogens that affect soybean crop yield. Soybean sudden death syndrome (SDS), caused by several distinct species of Fusarium, produces significant yield losses in the leading soybean-producing countries in North and South America. Current management strategies for SDS are scarce since there are no highly resistant cultivars and only a few fungicide seed treatments are available. Because of this, innovative approaches for SDS management need to be developed. Here, we summarize recently explored strategies based on plant nutrition, biological control, priming of plant defenses, host-induced gene silencing, and the development of new SDS-resistance cultivars using precision breeding techniques. Finally, sustainable management of SDS should also consider cultural control practices with minimal environmental impact. © 2021 Society of Chemical Industry.

Phaseocyclopentenones A and B, Phytotoxic Penta- and Tetrasubstituted Cyclopentenones Produced by Macrophomina phaseolina, the Causal Agent of Charcoal Rot of Soybean in Argentina.

Two new penta- and tetrasubstituted cyclopentenones, named phaseocyclopentenones A and B (1 and 2), together with guignardone A (3), were isolated from Macrophomina phaseolina cultures. The phytopathogenic fungus was isolated from infected soybean tissues showing charcoal rot symptoms in Argentina. Charcoal rot is a devastating disease considering that soybean is one of the main legumes cultivated in the world. Phaseocyclopentenones A and B were characterized by 1D and 2D 1H and 13C NMR spectroscopic and HRESIMS spectrometric data and chemical methods as 4-benzoyl-3,4,5-trihydroxy-2-phenylcyclopent-2-enone and 3,5-dihydroxy-2,4-diphenylcyclopent-2-enone, respectively. The relative configuration of phaseocyclopentenones A and B was assigned by 1H and NOESY NMR methods, while their absolute configurations were assigned by electronic circular dichroism methods. When assayed on a nonhost plant (Solanum lycopersicum L.) by the leaf puncture assay, phaseocyclopentenones A and B and guignardone A showed phytotoxic activity, while only 1 and 2 were toxic when tested on cuttings of the same plant. No phytotoxicity or antifungal activity was detected for the three compounds on the host plant soybean (Glycine max L.) and against some of its fungal pathogens, namely, Cercospora nicotianae and Colletotrichum truncatum, also isolated from infected soybean plants in Argentina.

Antifungal susceptibility profile and molecular identification of Cyp51C mutations in clinical and environmental isolates of Aspergillus flavus from Argentina.

BACKGROUND: The emergence of azole resistance in non-fumigatus Aspergillus strains is on the raise. OBJECTIVES: To study the susceptibility profiles and the molecular mechanisms of azole resistance of environmental and clinical strains of Aspergillus flavus from Argentina. METHODS: Thirty-five A flavus isolates (18 from soybean seeds and chickpea seeds and 17 from the clinic) were analysed for amphotericin B and azole resistance using the standard microbroth dilution method according to CLSI M38-A2 guidelines. Sequencing analysis of the cyp51 genes was conducted in those isolates displaying high MICs values to itraconazole, voriconazole and/or posaconazole. RESULTS: Among the environmental isolates, 33.3% of them showed high MIC values for at least one triazole whereas 23.5% of the clinical isolates displayed high MIC values for amphotericin B. Point mutations in the Cyp51C gene were recorded in most environmental isolates with non-wild-type MIC values. CONCLUSIONS: Susceptibility differences among environmental A flavus isolates might suggest the possibility of native resistance to certain triazole antifungals used in the clinic. To the best of our knowledge, this is the first report of antifungal screening of environmental strains of A flavus in soybean seeds and chickpea seeds from Argentina that showed increased resistance to voriconazole and itraconazole in comparison to clinical strains.

Role of Fungicide Applications on the Integrated Management of Wheat Stripe Rust.

First described in Europe in 1777, stripe rust (SR) caused by Puccinia striiformis Westend. f. sp. tritici Erikss (Pst) is one of the most important and destructive diseases of wheat worldwide. Until 2000, SR was mainly endemic to cooler regions, but since then, new aggressive strains have emerged, spread intercontinentally, and caused severe epidemics in warmer regions across the world. This has put SR as a disease that poses a threat to the world food security. At present, the preferred strategy for control of SR is the access to wheat cultivars with adequate levels of SR resistance. However, wheat breeding programs are not sufficiently advanced to cope with the recently emerged Pst strains. Under this scenario, foliar fungicide applications have become an important component of SR management, but information on the effects of fungicide applications on SR control and wheat cultivar yield response is scarce. This review seeks to provide an overview of the impact and role of fungicides on SR management. With focus on wheat management in the major wheat-growing regions of the world, the review addresses: (a) the efficacy of different fungicide active ingredients, optimal fungicide timing and number of applications in controlling SR, and (b) the impact of fungicide on wheat grain yield response. Inclusion of fungicides in an integrated crop management approach is discussed.

Draft genome sequence data of Cercospora kikuchii, a causal agent of Cercospora leaf blight and purple seed stain of soybeans.

Cercospora kikuchii (Tak. Matsumoto & Tomoy.) M.W. Gardner 1927 is an ascomycete fungal pathogen that causes Cercospora leaf blight and purple seed stain on soybean. Here, we report the first draft genome sequence and assembly of this pathogen. The C. kikuchii strain ARG_18_001 was isolated from soybean purple seed collected from San Pedro, Buenos Aires, Argentina, during the 2018 harvest. The genome was sequenced using a 2 × 150 bp paired-end method by Illumina NovaSeq 6000. The C. kikuchii protein-coding genes were predicted using FunGAP (Fungal Genome Annotation Pipeline). The draft genome assembly was 33.1 Mb in size with a GC-content of 53%. The gene prediction resulted in 14,856 gene models/14,721 protein coding genes. Genomic data of C. kikuchii presented here will be a useful resource for future studies of this pathosystem. The data can be accessed at GenBank under the accession number VTAY00000000 https://www.ncbi.nlm.nih.gov/nuccore/VTAY00000000.

Effect of potassium and manganese phosphites in the control of Pythium damping-off in soybean: a feasible alternative to fungicide seed treatments.

BACKGROUND: Use of fungicide seed treatments for control of soybean soilborne diseases such as Pythium damping-off has increased worldwide. However, emergence of Pythium strains resistant to metalaxyl-M has prompted the need for alternative technologies to fungicides for damping-off control. The use of phosphites (Phis) has been proposed as a method to control oomycetes, but their use as seed treatments in soybean is limited by the lack of information on their efficacy. The effect of potassium (K) and manganese (Mn) Phis (as seed treatments) in the control of Pythium damping-off in soybean was evaluated in vitro and in vivo. In vitro, treated seeds and a control were placed on potato dextrose agar and the damping-off severity caused by Pythium aphanidermatum (Edson) Fitzpatrick, Pythium irregulare Buisman, and Pythium ultimum Trow was assessed 5 days after incubation using an ordinal scale. In vivo, treated seeds and a control were planted in polystyrene pots and emergence was evaluated 21 days after planting. RESULTS: Analysis of the in vitro data using a multinomial generalized linear model showed that the probabilities of non-germinated, dead seeds ranged from 0.64 to 1.00 in the control and from 0 to 0.13 in the Phi treatments in each of the Pythium species. Probabilities of seed germination without or with damping-off symptoms were significantly higher for seeds treated with the Phi products than for the control. In the in vivo experiment, the Phi-based products increased seedling emergence by up to 29% on average compared with the untreated control. CONCLUSION: Mn and K Phis are feasible alternatives as seed treatments to control Pythium damping-off in soybean. This study is the first, worldwide, to document the efficacy of K and Mn Phis in the control of soybean Pythium damping-off. © 2017 Society of Chemical Industry.