Fusaric acid mediates the assembly of disease-suppressive rhizosphere microbiota via induced shifts in plant root exudates

Abstract

The plant health status is determined by the interplay of plant-pathogen-microbiota in the rhizosphere. Here, we investigate this tripartite system focusing on the pathogen Fusarium oxysporum f. sp. lycopersici (FOL) and tomato plants as a model system. First, we explore differences in tomato genotype resistance to FOL potentially associated with the differential recruitment of plant-protective rhizosphere taxa. Second, we show the production of fusaric acid by FOL to trigger systemic changes in the rhizosphere microbiota. Specifically, we show this molecule to have opposite effects on the recruitment of rhizosphere disease-suppressive taxa in the resistant and susceptible genotypes. Last, we elucidate that FOL and fusaric acid induce changes in the tomato root exudation with direct effects on the recruitment of specific disease-suppressive taxa. Our study unravels a mechanism mediating plant rhizosphere assembly and disease suppression by integrating plant physiological responses to microbial-mediated mechanisms in the rhizosphere.

Fig. 1. FOL and FA-induced tomato rhizosphere microbiota recruitment and Fusarium wilt disease resistance.

a Fusarium wilt disease severity of tomato plants grown in natural and sterile soils, respectively. S, susceptible cultivar D72; R, resistant cultivar Z19. b Contribution of microbiota-mediated disease resistance. c Schematic representation of the split-root system and rhizosphere transplant experiments to test the effect(s) of FOL on the tomato rhizosphere microbiota. d Fusarium wilt disease severity in the rhizosphere transplant experiment used to test the effect(s) of FOL on the tomato rhizosphere disease suppressiveness. Rhizosphere samples were collected from the systemic pot of FOL-inoculated plants (FOL.Sys) and from the non-inoculated control (FOL.Control). e FA content in the culture filtrate of FOL grown in potato dextrose broth. f FA content in the tomato rhizosphere in the split-root system used to test the effects of FOL on the rhizosphere microbiota. g Schematic representation of the split-root system and the rhizosphere transplant experiment used to test the effect(s) of FA on the rhizosphere disease suppressiveness. h Fusarium wilt disease severity in the rhizosphere transplant experiment used to test the effect(s) of FA on the rhizosphere disease suppressiveness. Rhizosphere samples were collected from the systemic pot of FA-treated plants (FA.Sys) and from the untreated control (FA.Control). FOL, F. oxysporum f. sp. lycopersici; FA, fusaric acid; Sys, systemic. In box plots, the center line represents the median, box edges delimit lower and upper quartiles and whiskers show the highest and lowest values. For (a, b, d–f, h), data are shown as mean ± SEM (n = 4). Different letters represent significant differences between treatments (Tukey’s HSD test, P < 0.05). ns, non-significant. P values were determined through two-sided Welch’s t tests. The tomato plant image was created with BioRender.com. Source data are provided as a Source Data file.

Fig. 2. FOL infection and FA alter the tomato rhizosphere bacterial community composition.

a Effects of FOL on the β-diversity of bacterial communities in the tomato rhizosphere of susceptible (S) and resistant (R) cultivars. b Comparison of bacterial OTUs altered due to FOL in the two tomato cultivars. c Effects of FA on the β-diversity of bacterial communities in the tomato rhizosphere of susceptible and resistant cultivars. d Comparison of bacterial OTUs altered due to FA in the two tomato cultivars. For (b) and (d), data are shown as the log₂-fold differences between FOL.Sys vs. FOL.Control (or FA.Sys vs. FA.Control) for each cultivar (two-sided Wald test with P values Benjamini-Hochberg adjusted). The size of each circle corresponds to its mean relative abundance (RA) across all samples. The color of each circle represents whether the taxon is altered by FOL (or FA) in only one cultivar or in both cultivars. FOL, F. oxysporum f. sp. lycopersici; FA, fusaric acid; Sys, systemic. Source data are provided as a Source Data file.

Fig. 3. Effects of bacterial isolates and distinct SynComs on Fusarium wilt disease suppression.

a Maximum likelihood phylogenetic tree based on the 16 S rRNA gene sequences of bacterial isolates. The sequence similarity of each Flavobacterium, Arthrobacter, Streptomyces, Lysobacter, Sphingobium and Sphingomonas spp. isolates with OTU3288, OTU1254, OTU1313, OTU3848, OTU4099 and OTU1379, respectively, is provided between parentheses. The NCBI database accession numbers of the closest phylogenetic relatives of isolates having the highest sequence similarity with these OTUs (in red color) are provided in brackets. Photos on the right panel show the in vitro antagonistic activity of selected isolates against FOL on potato dextrose agar. b Schematic representation of the pot experiment used to test the suppressive potential of bacterial isolates and SynComs on pathogen FOL. c Effects of individual bacterial isolates on Fusarium wilt disease severity. S, susceptible cultivar D72; R, resistant cultivar Z19. d Effects of the SynComs on Fusarium wilt disease severity. e Effects of bacterial-induced systemic resistance against Fusarium wilt disease. f Relative expression levels of defense-related genes in tomato roots. For each tomato cultivar, results are normalized to tomato ACTIN gene and expressed relative to those detected in non-inoculated plants before FOL infection—set as an arbitrary value of 1 (dashed line). FOL, F. oxysporum f. sp. lycopersici; SynCom, synthetic community. For (c), each dot represents one bacterial isolate (n = 4 for each isolate). Solid and open dots represent isolates with significant and non-significant disease suppressive effects (as compared with the non-inoculated control treatment). Blue dots represent the isolates Fl79, Ar03, St81, Ly56, Sb87, or Sm12. For (d–f), data are shown as mean ± SEM (n = 4). Different letters represent significant differences between treatments (Tukey’s HSD test; P < 0.05). The tomato plant image was created with BioRender.com. Source data are provided as a Source Data file.

Fig. 4. FOL and FA modulate the root colonization by Sphingomonas sp.

a Schematic representation of the split-root system used to test the root colonization by bacterial isolates and SynComs. b Effects of FOL infection on the root colonization by each bacterial isolate in the SynComs. S, susceptible cultivar D72; R, resistant cultivar Z19. c Effect of exogenous FA amendment on the abundances of Sm12 and Sphingomonas sp. in the rhizosphere of tomato. d Effect of the FOL wild-type (WT) and Δfub1 mutant on the abundances of Sm12 and Sphingomonas sp. in the rhizosphere of tomato. FOL, F. oxysporum f. sp. lycopersici; FA, fusaric acid; SynCom, synthetic community; CFU, colony forming units. For (b–d), data are shown as mean ± SEM (n = 4). Different letters represent significant differences between treatments (Tukey’s HSD test; P < 0.05). The tomato plant image was created with BioRender.com. Source data are provided as a Source Data file.

Fig. 5. Effect of FOL inoculation on tomato root exudates and root colonization by Sphingomonas sp. Sm12.

a Effects of tomato root exudates on the growth and biofilm formation of Sm12. Tomato root exudates were collected from the systemic pot of FOL-inoculated plants (FOL.Sys) and from the non-inoculated control (FOL.Control). S, susceptible cultivar D72; R, resistant cultivar Z19. b Orthogonal partial least squares-discriminant analysis (OPLS-DA) of metabolites in tomato root exudates. c Heatmap depicting the distinct metabolites in tomato root exudates between treatments. *indicates significant altered metabolites with variable importance of projection > 1, log₂ fold change > 1 and Benjamini-Hochberg adjusted P < 0.01 (two-sided Wald test). d Effects of specific metabolites on the growth and biofilm formation of Sm12. e Conceptual diagram of the mechanisms by which FOL-derived FA modulates the disease suppressive status of the tomato rhizosphere microbiota. In brief, FOL produces FA (1), which differentially alters the tomato root exudates in susceptible and resistant cultivars (2). These differences in root exudate metabolites impact the root colonization by disease-suppressive bacterial taxa (e.g., Sphingomonas sp.) (3), and thus modulate the status of plant health (4). Importantly, the root colonization by these bacteria were stimulated in the resistant cultivar Z19 but inhibited in the susceptible cultivar D72. These disease-suppressive bacteria act via induced systemic resistance (5). HA, 2-Hydroxyglutaric acid; FOL, F. oxysporum f. sp. lycopersici; FA, fusaric acid; OD, optical density. For (a, d), data are shown as mean ± SEM (n = 4). Different letters represent significant differences between treatments (Tukey’s HSD test; P < 0.05). The tomato plant image was created with BioRender.com. Source data are provided as a Source Data file.