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. 2025 Oct 1;41(5):595–606. doi: 10.5423/PPJ.OA.06.2025.0074

Dimethyl Sulfoxide Suppresses Fusarium graminearum Pathogenicity by Modulating Key Life Cycle Traits

Taiying Li 1,2, Seunghyun Lee 2, Taewon Choi 2, Yurim Lim 2, Tianling Ma 1,3, Jungkwan Lee 2,*
PMCID: PMC12488374  PMID: 41033672

Abstract

Dimethyl sulfoxide (DMSO) is widely recognized for its versatile solvent properties, and for its role as a cryoprotectant in the preservation of cell and microorganism. Despite its extensive use across various fields, its impact on plant pathogens has received comparatively less attention in existing literature. This study focuses on investigating the effects of DMSO on Fusarium graminearum, a fungal pathogen that affects grains, through both bioinformatic and biological experiments. Our findings demonstrate that, although DMSO induces mycotoxin production in F. graminearum in vitro, it significantly reduces production and maturation of perithecia and pigmentation. Additionally, DMSO supplementation inhibits mycelial growth and conidial germination, potentially contributing to reduced pathogenicity on wheat coleoptiles. These results highlight DMSO’s potential influence on plant pathogenic fungi beyond F. graminearum and may provide valuable insights for future research.

Keywords: DMSO, Fusarium graminearum; mycelial growth; pathogenicity; sexual reproduction

Dimethyl sulfoxide (DMSO), an organosulfur compound, combines broad solubility (polar and nonpolar compatibility) with straightforward synthesis (Martin et al., 1967). Due to these properties, DMSO is commonly utilized as a solvent to dissolve a variety of materials in various experiments. Additionally, DMSO finds extensive application as a cryoprotectant for cells and microorganisms during freezing and storage (Pegg, 2015). Besides these well-known uses, research on the effects of DMSO has been carried out in many fields.

In the field of medicine, DMSO serves a dual role. Firstly, DMSO is frequently employed as a vehicle for substances such as antibacterial, antifungal, antiviral, and membrane penetration enhancement agents (Carraher et al., 2013; David, 1972; Hoang et al., 2021; Marren, 2011). Moreover, DMSO serves as a promoter for enhancing the synthesis of secondary metabolites such as tetracenomycin C, chloramphenicol, and thiostrepton in bacteria like Streptomyces spp. and Bacillus spp., renowned for their antimicrobial production capabilities (Chen et al., 2000). Secondly, it is utilized directly for therapeutic purposes. Previous studies have demonstrated that DMSO treatment exerts significant inhibitory effects on the growth of melanoma cells in both B16 mouse melanoma cells and human melanoma cells, albeit resulting in a notable increase in cell pigmentation (Huberman et al., 1979). Furthermore, it has been reported that DMSO inhibits the mycelial growth of pathogenic fungi such as Microsporum canis and Trichophyton mentagrophytes in human skin (Huberman et al., 1979; Nordenberg et al., 1986; Randhawa, 2006). However, some studies have reported that in Aspergillus flavus, the addition of DMSO increased gene expression and synthesis of aflatoxins, which are toxins endanger food safety and international trade (Costes et al., 2021).

In the field of plant protection, studies involving the plant pathogenic fungi Aspergillus niger and Botrytis cinerea have demonstrated that DMSO treatment inhibits their pigmentation, mycelial growth, and pathogenicity (Carley et al., 1967; Petruccelli et al., 2020). Although there are some studies on the effects of DMSO on plant pathogens, multiple literature sources indicate that most research focuses on animal cells. In recent years, DMSO-related studies have primarily concentrated on human pathogens and fungi associated with environmental issues, while research on the effects of DMSO on plant pathogens remains extremely limited (Sun et al., 2023; Toreno et al., 2024). Therefore, in this study, we aimed to investigate the effects of DMSO on Fusarium graminearum, which cause disease in grains.

Fusarium graminearum is the causal agent of Fusarium head blight in major cereal crops such as rice, wheat, barley, and maize. As a characters, initially F. graminearum exhibits whitish mycelium, then developing into a yellow-orange, and finally turning into carmine-red (Cambaza, 2018). It also produces mycotoxins, including deoxynivalenol (DON) and zearalenone (ZEA), which lead to mycotoxicosis in human and livestock (Leslie and Summerell, 2006). Additionally, as an ascomycetous fungus, F. graminearum can produce sexual progeny without contact with a sexual partner (Kim et al., 2012). When F. graminearum is exposed to unsuitable conditions, such as winter, it continuously develops into flask-shaped perithecia that are filled with asci. The tubular sacs of asci contain ascospores, which reach the top of the perithecium and are forcibly discharge into the air when environmental conditions are favorable and the asci mature. This process is a critical part of the disease cycle for F. graminearum that infects grains (Guenther and Trail, 2005; Trail, 2009; Trail et al., 2002).

In this study, we conducted transcriptomic analysis and biological experiments on F. graminearum, integrating transcriptomic data with biological experiments results to identify morphological and physiological changes induced by DMSO. Our results indicated that DMSO affects normal gene expression associated with mycelial growth, reproduction, and pigmentation in F. graminearum. These differential gene expressed genes (DEGs) are closely linked to reduced mycelial growth, inhibited conidial germination, suppressed sexual reproduction, and diminished pigmentation in F. graminearum.

Materials and Methods

Fungal strains and media

The fungal strain F. graminearum GZ03639 (Bowden and Leslie, 1999) was used for all of the experiments and maintained in 20% glycerol at −80°C. Various media, including potato dextrose agar (PDA), potato dextrose broth (PDB), minimal medium (MM), and complete medium (CM), were prepared according to the protocols outlined in the Fusarium laboratory manual (Leslie and Summerell, 2006). Additionally, conidia were induced on yeast malt agar (YMA), and a fertility test was conducted on carrot agar (CA). For the induction of ZEA, glucose yeast extract peptone (GYEP) (Li et al., 2022b) was utilized, while for induction of DON, agmatine (5 mM) was included in MM (AMM) (Son et al., 2011a).

Mycelia growth and conidial germination

Fusarium graminearum was cultured on MM, CM, PDA, and GYEP media supplemented with 0%, 0.5%, 1%, 1.5%, and 2% of DMSO at 25°C. Botrytis cinerea, Magnaporthe oryzae, and Fusarium fujikuroi were cultured on MM supplemented with gradient of DMSO. After 3 days, the mycelia growth of F. graminearum on each medium was measured. Conidial germination was conducted as described previously with slight modifications (Li et al., 2019). In brief, mycelia blocks cultivated on CM for 3 days were inoculated into CM broth medium and incubated at 200 rpm at 25°C for 3 days. The mycelia were harvested using filter paper and then cultured on YMA medium before being incubated under near-ultraviolet light (20 W, 500 lux) at 25°C. After 2 days, the produced conidia were harvested by filtration with miracloth and centrifugation at 13,000 rpm at 4°C for 10 min. Subsequently, 200 μL of conidial suspension (1 × 106 conidia/mL) was inoculated into 10 mL MM supplemented with 0%, 0.25%, 0.75%, and 1.5% DMSO. The total number of conidia and the number of germinated conidia were counted 4, 8, and 12 h after inoculation using light microscopy.

Fertility test

Three-day-old mycelial plugs of F. graminearum were inoculated onto CA plates supplemented with 0%, 0.5%, 1%, 1.5%, and 2% DMSO, and then incubated at 25°C for 4 to 5 days in darkness for self-fertilization. The mycelia grown on the CA plates were removed using 1 mL of 2.5% Tween-20 solution. Subsequently, the plates were incubated under near-ultraviolet light at 25°C for 9 days to observe production of perithecia and asci (Lee et al., 2011). For asci observation, the perithecia were dissected in a drop of 20% glycerol on a glass slide, and the asci were flattened under a cover glass (Min et al., 2010).

Mycotoxin production

The production of DON was evaluated as described (He et al., 2007) with some modification. Briefly, conidia (1 × 104 conidia/mL) were cultured in 20 ml of AMM with or without 1.5% DMSO for 7 days at 25°C under stationary conditions. Subsequently, the culture solution was filtered through miracloth and then filtered again using 0.2 μm syringe filter. DON was extracted from 150 μL of culture filtrates by mixing with 250 μl of an ethyl acetate-methanol solution (4:1, v/v). Then 100 μL of supernatant was used to determine the concentration of DON, and DON concentrations were measured using enzyme-linked immunosorbent assay kit (CUSABIO, College Park, MD, USA) following the manufacturer’s instructions.

For production of ZEA, conidia (1 × 105 conidia/mL) were cultured in 20 mL of GYEP with or without 1.5% DMSO for 7 days at 200 rpm and 25°C. The mycelia were then removed using filter paper and filtered through a 0.2 μm syringe filter. The filtrate was mixed with n-hexane (1:1, v/v) in a separating funnel and vigorously shaken to achieve defatting. Subsequently, the aqueous layer was separated, and an equivalent volume of ethyl acetated was added to the separating funnel. After thorough mixing, only the upper ethyl acetate layer was collected into a round-bottom flask. The extraction with ethyl acetate was repeated twice, and the ethyl acetate was dried at 70°C using a Heidolph Rotary Evaporator (Heidolph, Schwabach, Germany) and dissolved in 1 mL of methanol for use as a sample. The concentration of ZEA was determined using Zearalenone ELISA kit (Demeditec Diagnostics GmbH, Kiel, Germany) following the manufacturer’s instructions.

Pigmentation test

The pigmentation of F. graminearum was observed on solid and liquid media, including PDA (or PDB), GYEP, MM, and CM. For pigmentation on solid media, 3-day-old F. graminearum was cultured on media supplemented with 0%, 0.5%, 1%, 1.5%, and 2% of DMSO for 15 days at 25°C. For pigmentation in liquid media, conidia (1 × 105 conidia/mL) were cultured in media supplemented with 0%, 0.25%, 0.75%, and 1.5% of DMSO for 5 days at 25°C and 200 rpm.

Extraction of aurofusarin

The extraction of aurofusarin was conducted following a previously described with slightly modification (Merhej et al., 2012). Briefly, F. graminearum was cultured on PDA supplemented with or without 1.5% DMSO for 10 days at 25°C. The plate was then dried in a hood for 3 days and ground into a powder. Twenty grams of the powder were placed into a 15 mL tube, followed by the addition of 10 mL of chloroform. Aurofusarin was subsequently extracted by shaking the solution in a water bath at 40°C for 1 h.

RNA extraction and sequencing

Fusarium graminearum conidia (1 × 105 conidia/mL) were cultured in GYEP supplemented with or without 0.75% DMSO at 200 rpm and 25°C. After 48 h of incubation, total RNA was extracted using the Easy-spin total RNA Extraction Kit (iNtRON Biotechnology, Seongnam, Korea) following the manufacturer’s protocol. Whole transcriptomes were sequenced using Illumina HiSeq 2000 (Illumina, San Diego, CA, USA) at Macrogen (Seoul, Korea).

Quantitative real time PCR

For confirming the influences of DMSO on expression of pigmentation-related genes, conidia (1 × 105 conidia/mL) were cultured in PDB supplemented with or without 1.5% DMSO for 8 days at 200 rpm and 25°C. To examine changes in gene expression related to DON synthesis, conidia (1 × 105 conidia/mL) were cultured in MM with or without 1.5% DMSO for 48 h at 200 rpm and 25°C. Mycelia were harvested, ground under liquid nitrogen, and total RNA was extracted using Easy-spin total RNA Extraction Kit according to the manufacturer’s protocol. Then, cDNA was generated using the First Strand cDNA Synthesis Kit (TOYOBO Co., Ltd., Osaka, Japan) according to the manufacturer’s instructions. The synthesized cDNA samples from each treatment were diluted to 100 ng/μL using distilled water, and 2 μL of cDNA was used for quantitative real time PCR (qRT-PCR). The qRT-PCR was performed according to the previous study using SYBR Green as fluorescent dye (Liang et al., 2024), and relative transcription levels were normalized by reference gene, cyclophilin (CYP). All of the PCR primers used in this study are listed in Supplementary Table 1.

Virulence test

For confirming the effect of DMSO on the virulence of F. graminearum, coleoptile test was performed following previous studies with some modifications (Li et al., 2022b; Ma et al., 2022). Briefly, wheat seeds (Geumgangmil) were soaked in 70% ethanol for 1 min, then in 3% sodium hypochlorite for 3 min, and washed twice with distilled water. The sterilized wheat seeds were germinated on moist filter paper at 25°C. The top 2–3 mm of the coleoptiles were removed, and 2 μL of conidia suspension (1 × 105 conidia/mL in distilled water or 1.5% DMSO) was inoculated. The coleoptiles were then cultivated in a growth chamber at 25°C with 100% relative humidity and 12 h of light and darkness each. The effect of DMSO on the virulence of F. graminearum was assessed by measuring the length of the lesion on the diseased stem 10 days after inoculation. The lesions length was measured using the ImageJ program (National Institutes of Health, Bethesda, MD, USA).

Quantitation of intracellular reactive oxygen species

The level of intracellular reactive oxygen species (ROS) in F. graminearum following DMSO supplementation were measured using the OxiSelect Intracellular ROS Assay Kit (Cell Biolabs, Inc., San Diego, CA, USA) according to the manufacturer’s instructions, with slight modifications. Briefly, conidia of F. graminearum were cultured in 10 mL of MM supplemented with 0% or 1.5% DMSO for 48 h at 25°C and 200 rpm. Subsequently, equal weights of germinated mycelia were collected and washed twice with phosphate-buffered saline. The mycelia were then incubated with dichloro-dihydro-fluorescein diacetate for 40 min at 37°C and washed twice. Finally, the mycelia were lysed in lysis buffer for 5 min, and fluorescence was measured at 480 nm (excitation) and 530 nm (emission) using a SpectraMax Gemini XPS plate reader (Molecular Devices, Sunnyvale, CA, USA).

Statistical analysis

The statistical differences in the number of perithecia and virulence test were examined using parametric one-way analysis of variance and the Kruskal-Wallis test, respectively, in R software version 4.4.0 (R Foundation for Statistical Computing, Vienna, Austria). Additionally, statistical differences in the remaining tests were examined using t-test.

Results

Effect of DMSO on transcriptome

Transcriptome data were deposited in the Sequence Read Archive at the National Center for Biotechnology Information (NCBI) under the accession number PRJNA1135744. According to the cutoff criteria (|Log2 fold change| ≥ 2 and P-value ≤ 0.05), we found that among the 1,444 DEGs, 1,060 genes were up-regulated and 384 genes were down-regulated in the medium supplemented with DMSO compared to the medium without DMSO (Supplementary Fig. 1).

To classify the functions of the identified DEGs, gene ontology (GO) analysis was performed. Among these GO classifications, we selected the top seven GO classifications in the biological process (BP) and molecular function (MF) categories, and the top GO classification in cellular component (CC) category. In the BP category, the transcripts were enriched in carbohydrate metabolic, cellular amino acid metabolic, lipid metabolic, protein-chromophore linkage, transmembrane transport, methionine biosynthesis, and metabolic process. Additionally, most transcripts were up-regulated. In the CC category, the majority of transcripts were enriched in the integral component of membrane, with more down-regulated transcripts being counted. In the MF category, the top transcripts were enriched in oxidoreductase activity, transmembrane transporter activity, phosphopantetheine binding, N-acetyltransferase activity, FMN binding, nitronate monooxygenase activity, and ATPase activity. Except N-acetyltransferase activity and ATPase activity, most transcripts were up-regulated (Fig. 1). Moreover, when screening the transcription factors related to normal mycelial growth, perithecia production and maturation, and pigmentation of F. graminearum, the supplementation of DMSO also led to the abnormal expression of many genes involved in transcription factors (Son et al., 2011b) (Supplementary Fig. 2).

Fig. 1.

Fig. 1

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Top enriched gene ontology (GO) terms of differentially expressed genes (DEGs). Top enriched GO terms of the DEGs ranked by the number of genes. BP, biological process; CC, cellular component; MF, molecular function.

DMSO reduces normal mycelial growth, conidial germination, and sexual development

In each medium, supplemented with DMSO significantly reduced mycelial growth compared to that without DMSO, and these reductions were more pronounced as the concentration of DMSO increased in F. graminearum wild-type, F. graminearum field strains (GWS 16-4-7 and GWR12R5), and other phytopathogens (Fig. 2, Supplementary Fig. 3A and D). Similarly, DMSO supplementation also decreased conidial germination in F. graminearum, especially when concentration of DMSO reached 0.75% (Supplementary Fig. 4).

Fig. 2.

Fig. 2

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Dimethyl sulfoxide (DMSO) reduces Fusarium graminearum mycelial growth. F. graminearum was grown on complete medium (CM), minimal medium (MM), potato dextrose agar (PDA), and glucose yeast extract peptone (GYEP) medium, each supplemented with increasing concentrations of DMSO, at 25°C for 3 days. Error bars represent the standard deviation from three replicates. Asterisks indicate P-value (*P < 0.05, **P < 0.01, ***P < 0.001) after comparison with t-test.

In CA medium, supplementation with DMSO significantly delayed the maturation of perithecia compared to that without DMSO, and the delay became more significant as the concentration of DMSO increased. Additionally, treatment with DMSO also significantly decreased the number of perithecia produced (Fig. 3, Supplementary Fig. 5).

Fig. 3.

Fig. 3

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Dimethyl sulfoxide (DMSO) reduces Fusarium graminearum sexual reproduction. The fungus was inoculated onto carrot agar, supplemented with increasing concentration of DMSO for 4 to 5 days. Sexual reproduction was then induced by placing the cultures under a near-ultraviolet lamp at 25°C. The produced perithecia and asci rosettes were observed at 5, 7, and 9 days. White scale bars = 500 μm, Black scale bars = 25 μm.

DMSO induces mycotoxin but reduces pathogenicity

The addition of DMSO to the media significantly increased the production of mycotoxins, DON and ZEA (Fig. 4A). Furthermore, supplementation with DMSO significantly elevated intracellular ROS levels and markedly up-regulated the transcript levels of Tri5 and Tri6 in DMSO-treated F. graminearum (Supplementary Fig. 3B and C). Interestingly, when inoculated into coleoptiles, conidia suspended in distilled water or DMSO, the former significantly increased lesions length. However, there was no difference between conidia suspended in DMSO and the control groups (DW and DMSO) (Fig. 4B, Supplementary Fig. 6). Furthermore, when F. graminearum was treated with DMSO, genes encoding enzymes involved in scavenging hydroxyl radical, such as peroxisome and catalase, were significantly up-regulated (Table 1).

Fig. 4.

Fig. 4

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Effect of dimethyl sulfoxide (DMSO) on Fusarium graminearum pathogenicity. (A) The deoxynivalenol (DON) and zearalenone (ZEA) concentration of F. graminearum. The supernatant of each culture was used to analyze DON and ZEA concentrations. The strain was cultured in agmatine minimal medium and glucose yeast extract peptone for producing DON and ZEA, respectively, each supplemented with or without DMSO, for 7 days at 25°C. Error bars denote standard deviation (SD) from three replicates. Asterisks indicate P-values from t-test (**P < 0.01, ***P < 0.001). (B) Virulence of F. graminearum. Wheat coleoptiles were inoculated with 2 μL conidial suspension (1 × 105 conidia/mL in distilled water or 1.5% DMSO), and lesion length was measured at 10 dpi using ImageJ. Error bars denote SD from five replicates. Values with different letters are significantly different according to Tukey’s test (P < 0.05).

Table 1.

Differentially expressed genes from RNA-seq

Genes DEGs (0.75% DMSO/0% DMSO) P-value
Peroxisome related genes
 FGSG_00308 2.165 0.035
 FGSG_00622 3.489 0.003
 FGSG_00840 3.600 0.006
 FGSG_01419 2.587 0.036
 FGSG_03071 5.590 <0.001
 FGSG_03154 3.316 0.014
 FGSG_05695 47.699 <0.001
 FGSG_07078 5.307 <0.001
 FGSG_09979 2.599 0.032
 FGSG_10853 5.975 <0.001
 FGSG_11196 2.615 0.024
 FGSG_12728 2.626 0.024
Catalase related genes
 FGSG_06554 11.338 <0.001
 FGSG_06596 2.26558 <0.001
 FGSG_06733 3.806505 0.002
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DEG, differentially expressed gene; DMSO, dimethyl sulfoxide.

DMSO reduces pigmentation

Pigmentation of F. graminearum was reduced, both on solid and in liquid media with supplemented with DMSO (Fig. 5, Supplementary Fig. 7). Furthermore, the addition of DMSO significantly up-regulate the transcription of the pks12 gene involved in aurofusarin biosynthesis, while the expression of other downstream genes was significantly down-regulated. Additionally, the color of aurofusarin extracted from the culture of F. graminearum noticeably faded in the samples containing DMSO (Fig. 6).

Fig. 5.

Fig. 5

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Dimethyl sulfoxide (DMSO) reduces Fusarium graminearum pigmentation. F. graminearum was grown on complete medium (CM), minimal medium (MM), potato dextrose agar (PDA), and glucose yeast extract peptone (GYEP) medium, each supplemented with increasing concentrations of DMSO, at 25°C for 15 days.

Fig. 6.

Fig. 6

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Dimethyl sulfoxide (DMSO) affects gene expression in aurofusarin biosynthesis. The color gradation from blue to red represents log2-fold change (1.5% DMSO vs. 0% DMSO), ranging from a minimum of −3 to maximum 3. Yellow stars represent significantly different values according to t-test. The aurofusarin (center) was extracted from Fusarium graminearum cultured on potato dextrose agar supplemented with or without 1.5% DMSO for 10 days at 25°C.

Discussion

Transcriptome analyses showed that the supplementation of DMSO led to the down-regulation of many genes related to normal mycelia growth of F. graminearum. These genes included not only transcription factors tightly related to mycelial growth but also transcripts related to the integral component of membrane, as revealed by GO analysis (Fig. 1, Supplementary Fig. 2). In fungi, the cell membrane is one of the most important components, and its integrity not only protects the fungus from environment stresses but also serves as the site for many transports, thereby controlling the movement of substances into and out of cell (Gooday, 1995; Sant et al., 2016). Due to these effects, many recent drugs or biological control strategy have targeted the integrity of cell membranes to inhibit fungal growth or kill fungi (Gao et al., 2023; Müller et al., 2013; Ostrosky-Zeichner et al., 2010; Shor and Perlin, 2015). Some studies on phytopathogens have also shown that gene defects related to cell membrane integrity in pathogenic fungi can lead to mycelial growth defects (Li et al., 2022a, 2022b; Ma et al., 2022).

Additionally, GO analysis results showed that DMSO treatment can also down-regulate more transcripts related to ATPase activity (Fig. 1). Similar to the integrity of cell membranes, fungal plasma membrane ATPase is crucial to cell physiology, with one of its main functions being nutrient uptake, which is closely associated with normal vegetative growth (Manzoor, 2016). Previous studies have shown that mycelial growth of Aspergillus nidulans treated with the ATPase inhibitor bafilomycin was significantly inhibited (Melin et al., 1999; Schachtschabel et al., 2012). Moreover, in Phytophthora parasitica which belong to Oomycetes, silencing the gene encoding plasma membrane ATPase resulted in the inhibition of zoospore germination (Zhang et al., 2012). Similar results were also found in our conidial germination test of DMSO-supplemented F. graminearum (Supplementary Fig. 4). Therefore, combining previous studies and our GO analysis results, the supplementation of DMSO to F. graminearum negatively impacts the expression of transcripts associated with cell membrane integrity and ATPase activity. These gene-level negative effects are consistent with our observed reductions in mycelial growth and conidial germination in F. graminearum (Fig. 2), indicating that DMSO adversely affects its normal vegetative growth.

Co-existing with the host plant, head blight disease initiates with the landing of airborne spores of F. graminearum on flowering spikelet, followed by spore germination and eventual entry into the plant (Bushnell et al., 2003). After entering host cells, the fungus grows intercellularly, spreading through the xylem and pith (Guenther and Trail, 2005; Jansen et al., 2005; Trail, 2009). It is evident that the germination of fungal spores and the growth of mycelia are crucial for the infection of host plant in F. graminearum. However, in the whole life cycle of F. graminearum, there are also diseases caused by ascospores produced during sexual development of F. graminearum (Trail, 2009). Previous studies have shown that in field trials, compared to controls, eliminating the sexual stage of F. graminearum resulted in a substantial reduction in disease (Desjardins et al., 2006). Therefore, in this study, we also evaluated the effect of DMSO on the sexual reproduction of F. graminearum and found that the supplementation of DMSO led to the abnormal expression of a large number of genes encoding sexual reproduction-related transcription factors (Supplementary Fig. 2). Additionally, DMSO supplementation significantly reduced perithecia production but also delayed the maturation of the asci (Fig. 3, Supplementary Fig. 5).

Pathogenicity is one of the most important characteristics of F. graminearum, which causes serious grain yield loss. Therefore, in this study, we determined the effect of DMSO on pathogenicity by inoculating F. graminearum on wheat coleoptile. The results showed that DMSO resulted in the reduction of lesions caused by F. graminearum (Fig. 4B, Supplementary Fig. 6). Similar to previous studies of DMSO in animal cells or human pathogenic fungi (Costes et al., 2021; Huberman et al., 1979), two diametrically opposed results also emerged in our study. That is, although DMSO supplementation significantly reduced the pathogenicity of F. graminearum in the coleoptile experiment, it was interesting to note that DMSO induced mycotoxin production, including DON, by F. graminearum in vitro (Fig. 4A).

The mycotoxin DON, produced by F. graminearum not only affects crop yields but also causes acute vomiting and appetite suppression in animals that eat contaminated crops (Leplat et al., 2013; Warth et al., 2013). DON biosynthesis is triggered by a number of factors, one of which is oxidative stress experienced by pathogenic fungi during infection (Chen et al., 2019; Jung et al., 2018). Previous studies have also shown that DMSO can lead to an increase in ROS levels in cells (Dludla et al., 2018; Mannan et al., 2010). Accordingly, we assessed intracellular ROS accumulation in DMSO-supplemented F. graminearum and observed that DMSO significantly elevated ROS levels. Our RNA-seq analysis further revealed that DMSO supplementation significantly up-regulates genes associated with ROS scavenging enzymes, such as those for peroxisomes and catalases (Table 1). Additionally, DMSO supplementation up-regulated the expression of Tri5 and Tri6 genes, both of which are involved in DON biosynthesis, suggesting that the increased intracellular ROS induced by DMSO contributes to elevated DON production in F. graminearum. Moreover, we monitored the expression of genes related to the biosynthesis pathway of aurofusarin, one of the important pigments produced by F. graminearum. In the biosynthetic pathway, based on RNA-seq analysis, we further confirmed through qRT-PCR that DMSO supplementation significantly up-regulated the pks12 gene expression, while most downstream genes are significantly down-regulated. Correspondingly, consistent with gene expression results, the DMSO-treated group produced less aurofusarin compared to the control group (Fig. 6).

Pigmentation is also an important characteristic of F. graminearum, which changes color from yellow to orange to red as the fungus ages (Kim et al., 2006). There are still many unanswered questions about the relationship between changes in mold pigmentation patterns and F. graminearum lifecycle. Previous studies have identified several pigments produced by F. graminearum, such as aurofusarin, rubrofusarin, perithecial melanin, and torulene (Ashley et al., 1937; Avalos et al., 2017; Cambaza, 2018; Frandsen et al., 2016). Additionally, although the pigments produced by F. graminearum, especially aurofusarin, are less toxic than mycotoxin, many previous studies have demonstrated a connection between mycotoxins and pigments. For instance, histone H3 lysine 4-methylation is important for the transcription of both biosynthetic genes of DON and aurofusarin (Liu et al., 2015), and aurofusarin shift the gut microbes to a more harmful constitution, causing illness by inhibiting probiotic bacteria (Sondergaard et al., 2016).

Taken together, this study elucidates effects of DMSO on plant pathogenic fungus F. graminearum, revealing that DMSO reduces production of perithecia and pigmentation and delays ascospore maturation. Furthermore, DMSO enhances mycotoxin production in vitro, it decreases the pathogenicity of F. graminearum on wheat coleoptiles, likely due to its inhibition of mycelial growth and conidial germination. The purpose of this study is not to advocate DMSO as a substitute for chemical fungicide but to enhance our understanding of its potential effects on plant pathogenic fungi beyond F. graminearum. By providing evidence that disrupting the normal life cycle of pathogens can reduce plant disease incidence, this study provides valuable insights for future research.

Footnotes

Conflicts of Interest

No potential conflict of interest relevant to this article was reported.

Acknowledgments

This work was supported by Basic Science Research Program through the National Research Foundation of Korea funded by the Ministry of Education (2023R1A2C1005998), Biomaterials Specialized Graduate Program through the Korea Environmental Industry & Technology Institute (KEITI) funded by the Ministry of Environment (MOE), and National Science Foundation of China (32202241).

Electronic Supplementary Material

Supplementary materials are available at The Plant Pathology Journal website (http://www.ppjonline.org/).

References

  1. Ashley J. N., Hobbs B. C., Raistrick H. Studies in the biochemistry of micro-organisms: the crystalline colouring matters of Fusarium culmorum (W. G. Smith) Sacc. and related forms. Biochem. J. 1937;31:385–397. doi: 10.1042/bj0310385. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Avalos J., Pardo-Medina J., Parra-Rivero O., Ruger-Herreros M., Rodríguez-Ortiz R., Hornero-Méndez D., Limón M. C. Carotenoid biosynthesis in Fusarium. J. Fungi. 2017;3:39. doi: 10.3390/jof3030039. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bowden R. L., Leslie J. F. Sexual recombination in Gibberella zeae. Phytopathology. 1999;89:182–188. doi: 10.1094/PHYTO.1999.89.2.182. [DOI] [PubMed] [Google Scholar]
  4. Bushnell W. R., Hazen B. E., Pritsch C. Histology and physiology of Fusarium head blight. In: Leonard K. J., Bushnell W. R., editors. Fusarium head blight of wheat and barley. APS Press; St Paul, MN, USA: 2003. pp. 44–83. [Google Scholar]
  5. Cambaza E. Comprehensive description of Fusarium graminearum pigments and related compounds. Foods. 2018;7:165. doi: 10.3390/foods7100165. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Carley H. E., Watson R. D., Huber D. M. Inhibition of pigmentation in Aspergillus niger by dimethylsulfoxide. Can. J. Bot. 1967;45:1451–1453. [Google Scholar]
  7. Carraher C. E., Jr, Barot G., Shahi K., Roner M. R. Influence of DMSO on the inhibition of various cancer cells by water-soluble organotin polyethers. J. Chin. Adv. Mater. Soc. 2013;1:294–304. [Google Scholar]
  8. Chen G., Wang G. Y., Li X., Waters B., Davies J. Enhanced production of microbial metabolites in the presence of dimethyl sulfoxide. J. Antibiot. 2000;53:1145–1153. doi: 10.7164/antibiotics.53.1145. [DOI] [PubMed] [Google Scholar]
  9. Chen Y., Kistler H. C., Ma Z. Fusarium graminearum trichothecene mycotoxins: biosynthesis, regulation, and management. Annu. Rev. Phytopathol. 2019;57:15–39. doi: 10.1146/annurev-phyto-082718-100318. [DOI] [PubMed] [Google Scholar]
  10. Costes L. H., Lippi Y., Naylies C., Jamin E. L., Genthon C., Bailly S., Oswald I. P., Bailly J.-D., Puel O. The solvent dimethyl sulfoxide affects physiology, transcriptome and secondary metabolism of Aspergillus flavus. J. Fungi. 2021;7:1055. doi: 10.3390/jof7121055. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. David N. A. The pharmacology of dimethyl sulfoxide. Annu. Rev. Pharmacol. 1972;12:353–374. doi: 10.1146/annurev.pa.12.040172.002033. [DOI] [PubMed] [Google Scholar]
  12. Desjardins A. E., Plattner R. D., Shaner G., Brown D. W., Buechley G., Proctor R. H., Turgeon B. G. Field release of Gibberella zeae genetically modified to lack ascospores. Proceedings of the 2006 National Fusarium Head Blight Forum; Lexington, KY, USA: University of Kentucky; 2006. pp. 39–44. [Google Scholar]
  13. Dludla P. V., Jack B., Viraragavan A., Pheiffer C., Johnson R., Louw J., Muller C. J. F. A dose-dependent effect of dimethyl sulfoxide on lipid content, cell viability and oxidative stress in 3T3-L1 adipocytes. Toxicol. Rep. 2018;5:1014–1020. doi: 10.1016/j.toxrep.2018.10.002. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Frandsen R. J. N., Rasmussen S. A., Knudsen P. B., Uhlig S., Petersen D., Lysøe E., Gotfredsen C. H., Giese H., Larsen T. O. Black perithecial pigmentation in Fusarium species is due to the accumulation of 5-deoxybostrycoidin-based melanin. Sci. Rep. 2016;6:26206. doi: 10.1038/srep26206. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gao M., Abdallah M. F., Song M., Xu Y., Sun D., Lu P., Wang J. Novel endophytic Pseudescherichia sp. GSE25 strain significantly controls Fusarium graminearum and reduces deoxynivalenol in wheat. Toxins. 2023;15:702. doi: 10.3390/toxins15120702. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Gooday G. Cell membrane. In: Gow N. A. R., Gadd G. M., editors. The growing fungus. Springer; Dordrecht, Netherlands: 1995. pp. 63–74. [Google Scholar]
  17. Guenther J. C., Trail F. The development and differentiation of Gibberella zeae (anamorph: Fusarium graminearum) during colonization of wheat. Mycologia. 2005;97:229–237. doi: 10.3852/mycologia.97.1.229. [DOI] [PubMed] [Google Scholar]
  18. He J., Yang R., Zhou T., Tsao R., Young J. C., Zhu H., Li X.-Z., Boland G. J. Purification of deoxynivalenol from Fusarium graminearum rice culture and mouldy corn by high-speed counter-current chromatography. J. Chromatogr. A. 2007;1151:187–192. doi: 10.1016/j.chroma.2007.01.112. [DOI] [PubMed] [Google Scholar]
  19. Hoang C., Nguyen A. K., Nguyen T. Q., Fang W., Han B., Hoang B. X., Tran H. D. Application of dimethyl sulfoxide as a therapeutic agent and drug vehicle for eye diseases. J. Ocul. Pharmacol. Ther. 2021;37:441–451. doi: 10.1089/jop.2021.0043. [DOI] [PubMed] [Google Scholar]
  20. Huberman E., Heckman C., Langenbach R. Stimulation of differentiated functions in human melanoma cells by tumor-promoting agents and dimethyl sulfoxide. Cancer Res. 1979;39:2618–2624. [PubMed] [Google Scholar]
  21. Jansen C., von Wettstein D., Schäfer W., Kogel K.-H., Felk A., Maier F. J. Infection patterns in barley and wheat spikes inoculated with wild-type and trichodiene synthase gene disrupted Fusarium graminearum. Proc. Natl. Acad. Sci. U. S. A. 2005;102:16892–16897. doi: 10.1073/pnas.0508467102. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Jung B., Park J., Kim N., Li T., Kim S., Bartley L. E., Kim J., Kim I., Kang Y., Yun K., Choi Y., Lee H.-H., Ji S., Lee K. S., Kim B. Y., Shon J. C., Kim W. C., Liu K.-H., Yoon D., Kim S., Seo Y.-S., Lee J. Cooperative interactions between seed-borne bacterial and air-borne fungal pathogens on rice. Nat. Commun. 2018;9:31. doi: 10.1038/s41467-017-02430-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kim H.-K., Cho E. J., Lee S., Lee Y.-S., Yun S.-H. Functional analyses of individual mating-type transcripts at MAT loci in Fusarium graminearum and Fusarium asiaticum. FEMS Microbiol. Lett. 2012;337:89–96. doi: 10.1111/1574-6968.12012. [DOI] [PubMed] [Google Scholar]
  24. Kim J.-E., Jin J., Kim H., Kim J.-C., Yun S.-H., Lee Y.-W. GIP2, a putative transcription factor that regulates the aurofusarin biosynthetic gene cluster in Gibberella zeae. Appl. Environ. Microbiol. 2006;72:1645–1652. doi: 10.1128/AEM.72.2.1645-1652.2006. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Lee S., Son H., Lee J., Lee Y.-R., Lee Y.-W. A putative ABC transporter gene, ZRA1, is required for zearalenone production in Gibberella zeae. Curr. Genet. 2011;57:343–351. doi: 10.1007/s00294-011-0352-4. [DOI] [PubMed] [Google Scholar]
  26. Leplat J., Friberg H., Abid M., Steinberg C. Survival of Fusarium graminearum, the causal agent of Fusarium head blight: a review. Agron. Sustain. Dev. 2013;33:97–111. [Google Scholar]
  27. Leslie J. F., Summerell B. A. The Fusarium laboratory manual. Blackwell Publishing; Ames, IA, USA: 2006. p. 388. [Google Scholar]
  28. Li J., Yang S., Li D., Peng L., Fan G., Pan S. The plasma membrane H+-ATPase is critical for cell growth and pathogenicity in Penicillium digitatum. Appl. Microbiol. Biotechnol. 2022a;106:5123–5136. doi: 10.1007/s00253-022-12036-4. [DOI] [PubMed] [Google Scholar]
  29. Li T., Jung B., Park S.-Y., Lee J. Survival factor gene FgSvf1 is required for normal growth and stress resistance in Fusarium graminearum. Plant Pathol. J. 2019;35:393–405. doi: 10.5423/PPJ.OA.03.2019.0070. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Li T., Kim D., Lee J. NADPH oxidase gene, FgNoxD, plays a critical role in development and virulence in Fusarium graminearum. Front. Microbiol. 2022b;13:822682. doi: 10.3389/fmicb.2022.822682. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Liang D., Jiang Y., Zhang Y., Mao C., Ma T., Zhang C. The comparative genomics of Botryosphaeriaceae suggests gene families of Botryosphaeria dothidea related to pathogenicity on Chinese Hickory Tree. J. Fungi. 2024;10:299. doi: 10.3390/jof10040299. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Liu Y., Liu N., Yin Y., Chen Y., Jiang J., Ma Z. Histone H3K4 methylation regulates hyphal growth, secondary metabolism and multiple stress responses in Fusarium graminearum. Environ. Microbiol. 2015;17:4615–4630. doi: 10.1111/1462-2920.12993. [DOI] [PubMed] [Google Scholar]
  33. Ma T., Li Y., Lou Y., Shi J., Sun K., Ma Z., Yan L., Yin Y. The drug H+ antiporter FgQdr2 is essential for multiple drug resistance, ion homeostasis, and pathogenicity in Fusarium graminearum. J. Fungi. 2022;8:1009. doi: 10.3390/jof8101009. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Mannan A., Liu C., Arsenault P. R., Towler M. J., Vail D. R., Lorence A., Weathers P. J. DMSO triggers the generation of ROS leading to an increase in artemisinin and dihydroartemisinic acid in Artemisia annua shoot cultures. Plant Cell Rep. 2010;29:143–152. doi: 10.1007/s00299-009-0807-y. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Manzoor N. Plasma membrane ATPase: potential target for antifungal drug therapy. In: Chakraborti S., Dhalla N., editors. Regulation of Ca2+-ATPases, V-ATPases and F-ATPases: Advances in biochemistry in health and disease. Vol. 14. Springer; Cham, Switzerland: 2016. pp. 519–530. [Google Scholar]
  36. Marren K. Dimethyl sulfoxide: an effective penetration enhancer for topical administration of NSAIDs. Phys. Sportsmed. 2011;39:75–82. doi: 10.3810/psm.2011.09.1923. [DOI] [PubMed] [Google Scholar]
  37. Martin H. D., Weise A., Niclas H.-J. The solvent dimethyl sulfoxide. Angew. Chem. Int. Ed. Engl. 1967;6:318–334. doi: 10.1002/anie.196703181. [DOI] [PubMed] [Google Scholar]
  38. Melin P., Schnürer J., Wagner E. G. H. Changes in Aspergillus nidulans gene expression induced by bafilomycin, a Streptomyces-produced antibiotic. Microbiology. 1999;145:1115–1122. doi: 10.1099/13500872-145-5-1115. [DOI] [PubMed] [Google Scholar]
  39. Merhej J., Urban M., Dufresne M., Hammond-Kosack K. E., Richard-Forget F., Barreau C. The velvet gene, FgVe1, affects fungal development and positively regulates trichothecene biosynthesis and pathogenicity in Fusarium graminearum. Mol. Plant Pathol. 2012;13:363–374. doi: 10.1111/j.1364-3703.2011.00755.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Min K., Lee J., Kim J.-C., Kim S. G., Kim Y. H., Vogel S., Trail F., Lee Y.-W. A novel gene, ROA, is required for normal morphogenesis and discharge of ascospores in Gibberella zeae. Eukaryot. Cell. 2010;9:1495–1503. doi: 10.1128/EC.00083-10. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Müller C., Staudacher V., Krauss J., Giera M., Bracher F. A convenient cellular assay for the identification of the molecular target of ergosterol biosynthesis inhibitors and quantification of their effects on total ergosterol biosynthesis. Steroids. 2013;78:483–493. doi: 10.1016/j.steroids.2013.02.006. [DOI] [PubMed] [Google Scholar]
  42. Nordenberg J., Wasserman L., Beery E., Aloni D., Malik H., Stenzel K. H., Novogrodsky A. Growth inhibition of murine melanoma by butyric acid and dimethylsulfoxide. Exp. Cell Res. 1986;162:77–85. doi: 10.1016/0014-4827(86)90427-1. [DOI] [PubMed] [Google Scholar]
  43. Ostrosky-Zeichner L., Casadevall A., Galgiani J. N., Odds F. C., Rex J. H. An insight into the antifungal pipeline: selected new molecules and beyond. Nat. Rev. Drug Discov. 2010;9:719–727. doi: 10.1038/nrd3074. [DOI] [PubMed] [Google Scholar]
  44. Pegg D. E. Principles of cryopreservation. Methods Mol. Biol. 2015;1257:3–19. doi: 10.1007/978-1-4939-2193-5_1. [DOI] [PubMed] [Google Scholar]
  45. Petruccelli V., Brasili E., Varone L., Valletta A., Pasqua G. Antifungal activity of dimethyl sulfoxide against Botrytis cinerea and phytotoxicity on tomato and lettuce plants. Plant Biosyst. 2020;154:455–462. [Google Scholar]
  46. Randhawa M. A. The effect of dimethyl sulfoxide (DMSO) on the growth of dermatophytes. Jpn. J. Med. Mycol. 2006;47:313–318. doi: 10.3314/jjmm.47.313. [DOI] [PubMed] [Google Scholar]
  47. Sant D. G., Tupe S. G., Ramana C. V., Deshpande M. V. Fungal cell membrane-promising drug target for antifungal therapy. J. Appl. Microbiol. 2016;121:1498–1510. doi: 10.1111/jam.13301. [DOI] [PubMed] [Google Scholar]
  48. Schachtschabel D., Arentshorst M., Lagendijk E. L., Ram A. F. J. Vacuolar H+-ATPase plays a key role in cell wall biosynthesis of Aspergillus niger. Fungal Genet. Biol. 2012;49:284–293. doi: 10.1016/j.fgb.2011.12.008. [DOI] [PubMed] [Google Scholar]
  49. Shor E., Perlin D. S. Coping with stress and the emergence of multidrug resistance in fungi. PLoS Pathog. 2015;11:e1004668. doi: 10.1371/journal.ppat.1004668. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Son H., Lee J., Park A. R., Lee Y.-W. ATP citrate lyase is required for normal sexual and asexual development in Gibberella zeae. Fungal Genet. Biol. 2011a;48:408–417. doi: 10.1016/j.fgb.2011.01.002. [DOI] [PubMed] [Google Scholar]
  51. Son H., Seo Y.-S., Min K., Park A. R., Lee J., Jin J.-M., Lin Y., Cao P., Hong S.-Y., Kim E.-K., Lee S.-H., Cho A., Lee S., Kim M.-G., Kim Y., Kim J.-E., Kim J.-C., Choi G. J., Yun S.-H., Lim J. Y., Kim M., Lee Y.-H., Choi Y.-D., Lee Y.-W. A phenome-based functional analysis of transcription factors in the cereal head blight fungus, Fusarium graminearum. PLoS Pathog. 2011b;7:e1002310. doi: 10.1371/journal.ppat.1002310. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Sondergaard T. E., Fredborg M., Oppenhagen Christensen A.-M., Damsgaard S. K., Kramer N. F., Giese H., Sørensen J. L. Fast screening of antibacterial compounds from fusaria. Toxins. 2016;8:355. doi: 10.3390/toxins8120355. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Sun B., Paraskevopoulos G., Min J., Rossdeutcher R., Ghosh S., Quinn B., Lin M.-H., Sarkar D., Sukumaran D., Wang Y., Vávrová K., Lovell J. F., Zhang Y. Topical drug delivery of concentrated cabazitaxel in a α-topherol and DMSO solution. Adv. Sci. 2023;10:e2302658. doi: 10.1002/advs.202302658. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Toreno G., Zucconi L., Caneva G., Meloni P., Isola D. Recolonization dynamics of marble monuments after cleaning treatments: a nine-year follow-up study. Sci. Total Environ. 2024;912:169350. doi: 10.1016/j.scitotenv.2023.169350. [DOI] [PubMed] [Google Scholar]
  55. Trail F. For blighted waves of grain: Fusarium graminearum in the postgenomics era. Plant Physiol. 2009;149:103–110. doi: 10.1104/pp.108.129684. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Trail F., Xu H., Loranger R., Gadoury D. Physiological and environmental aspects of ascospore discharge in Gibberella zeae (anamorph Fusarium graminearum) Mycologia. 2002;94:181–189. [PubMed] [Google Scholar]
  57. Warth B., Sulyok M., Berthiller F., Schuhmacher R., Krska R. New insights into the human metabolism of the Fusarium mycotoxins deoxynivalenol and zearalenone. Toxicol. Lett. 2013;220:88–94. doi: 10.1016/j.toxlet.2013.04.012. [DOI] [PubMed] [Google Scholar]
  58. Zhang M., Meng Y., Wang Q., Liu D., Quan J., Hardham A. R., Shan W. PnPMA1, an atypical plasma membrane H+-ATPase, is required for zoospore development in Phytophthora parasitica. Fungal Biol. 2012;116:1013–1023. doi: 10.1016/j.funbio.2012.07.006. [DOI] [PubMed] [Google Scholar]

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Supplementary Materials

PPJ-OA-06-2025-0074-Supplementary-Fig-1.pdf (302.6KB, pdf)
PPJ-OA-06-2025-0074-Supplementary-Fig-2,3.pdf (572.9KB, pdf)
PPJ-OA-06-2025-0074-Supplementary-Fig-4,5.pdf (744.3KB, pdf)
PPJ-OA-06-2025-0074-Supplementary-Fig-6.pdf (378.5KB, pdf)
PPJ-OA-06-2025-0074-Supplementary-Fig-7.pdf (450.3KB, pdf)
PPJ-OA-06-2025-0074-Supplementary-Table-1.pdf (302.6KB, pdf)

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