Plant secondary metabolites that are released into the rhizosphere alter biotic and abiotic soil properties, which in turn affect the performance of other plants. How such plant-soil feedbacks affect agricultural productivity and food quality in crop rotations is unknown. Here, we assessed the impact of maize benzoxazinoids on the performance, yield and food quality of three winter wheat varieties in a two-year field experiment. Following maize cultivation, we detected benzoxazinoid-dependent chemical and microbial fingerprints in the soil. The chemical fingerprint was still visible during wheat growth, while the microbial fingerprint was no longer detected. Benzoxazinoid soil conditioning by wild-type maize led to increased wheat emergence, tillering, growth and biomass compared to soil conditioning by bx1 mutant plants. Weed cover remained unaffected, while insect damage decreased in a subset of varieties. Wheat yield was increased by over 4% without reduction in grain quality across variety. This improvement was directly associated with increased germination and tillering. Taken together, our experiments demonstrate that plant secondary metabolites can increase yield via plant-soil feedbacks under agronomically realistic conditions. If this phenomenon holds across different soils and environmental conditions, optimizing plant root exudation could be a powerful, genetically tractable strategy to enhance crop yields without additional inputs.

Arbuscular mycorrhiza fungi (AMF) are beneficial soil fungi that can promote the growth of their host plants. Accurate quantification of AMF in plant roots is important because the level of colonization is often indicative of the activity of these fungi. Root colonization is traditionally measured with microscopy methods which visualize fungal structures inside roots. Microscopy methods are labor-intensive, and results depend on the observer. In this study, we present a relative qPCR method to quantify AMF in which we normalized the AMF qPCR signal relative to a plant gene. First, we validated the primer pair AMG1F and AM1 in silico, and we show that these primers cover most AMF species present in plant roots without amplifying host DNA. Next, we compared the relative qPCR method with traditional microscopy based on a greenhouse experiment with Petunia plants that ranged from very high to very low levels of AMF root colonization. Finally, by sequencing the qPCR amplicons with MiSeq, we experimentally confirmed that the primer pair excludes plant DNA while amplifying mostly AMF. Most importantly, our relative qPCR approach was capable of discriminating quantitative differences in AMF root colonization and it strongly correlated (Spearman Rho = 0.875) with quantifications by traditional microscopy. Finally, we provide a balanced discussion about the strengths and weaknesses of microscopy and qPCR methods. In conclusion, the tested approach of relative qPCR presents a reliable alternative method to quantify AMF root colonization that is less operator-dependent than traditional microscopy and offers scalability to high-throughput analyses.

Plant secondary metabolites that are released into the rhizosphere alter biotic and abiotic soil properties, which in turn affect the performance of other plants. How this type of plant-soil feedback affects agricultural productivity and food quality in the field in the context of crop rotations is unknown. Here, we assessed the performance, yield and food quality of three winter wheat varieties growing in field plots whose soils had been conditioned by either wild type or benzoxazinoid-deficient bx1 maize mutant plants. Following maize cultivation, we detected benzoxazinoid-dependent chemical and microbial fingerprints in the soil. The benzoxazinoid fingerprint was still visible during wheat growth, but the microbial fingerprint was no longer detected. Wheat emergence, tillering, growth, and biomass increased in wild type conditioned soils compared to bx1 mutant conditioned soils. Weed cover was similar between soil conditioning treatments, but insect herbivore abundance decreased in benzoxazinoid-conditioned soils. Wheat yield was increased by over 4% without a reduction in grain quality in benzoxazinoid-conditioned soils. This improvement was directly associated with increased germination and tillering. Taken together, our experiments provide evidence that soil conditioning by plant secondary metabolite producing plants can increase yield via plant-soil feedbacks under agronomically realistic conditions. If this phenomenon holds true across different soils and environments, optimizing root exudation chemistry could be a powerful, genetically tractable strategy to enhance crop yields without additional inputs.

Plants exude specialized metabolites from their roots and these compounds are known to structure the root microbiome. However, the underlying mechanisms are poorly understood. We established a representative collection of maize root bacteria and tested their tolerance against benzoxazinoids, the dominant specialized and bioactive metabolites in the root exudates of maize plants. In vitro experiments revealed that benzoxazinoids inhibited bacterial growth in a strain- and compound-dependent manner. Tolerance against these selective antimicrobial compounds depended on bacterial cell wall structure. Further, we found that native root bacteria isolated from maize tolerated the benzoxazinoids better compared to non-host Arabidopsis bacteria. This finding suggests the adaptation of the root bacteria to the specialized metabolites of their host plant. Bacterial tolerance to 6-methoxy-benzoxazolin-2-one (MBOA), the most abundant and selective antimicrobial metabolite in the maize rhizosphere, correlated significantly with the abundance of these bacteria on benzoxazinoid-exuding maize roots. Thus, strain-dependent tolerance to benzoxazinoids largely explained the abundance pattern of bacteria on maize roots. Abundant bacteria generally tolerated MBOA, while low abundant root microbiome members were sensitive to this compound. Our findings reveal that tolerance to plant specialized metabolites is an important competence determinant for root colonization. We propose that bacterial tolerance to plant-secreted antimicrobial compounds is an underlying mechanism determining the structure of host-specific microbial communities.

IntroductionHarnessing positive plant–soil feedbacks via crop rotations is a promising strategy for sustainable agriculture. These feedbacks are often context‐dependent, and how soil heterogeneity explains this variation is unknown. Plants influence soil properties, including microbes, by exuding specialized metabolites. Benzoxazinoids, specialized metabolites released by cereals such as wheat and maize, can alter rhizosphere microbiota and performance of plants subsequently growing in the exposed soils and are thus an excellent model to study agriculturally relevant plant–soil feedbacks.Materials and MethodsTo understand local variation in soil properties on benzoxazinoid‐mediated plant–soil feedbacks, we conditioned plots with wild‐type maize and benzoxazinoid‐deficient bx1 mutants in a grid pattern across a field, and we then grew winter wheat in the following season. We determined accumulation of benzoxazinoids, root‐associated microbial communities, abiotic soil properties and wheat performance in each plot and then assessed their associations.ResultsWe detected a marked gradient in soil chemistry and microbiota across the field. This gradient resulted in significant differences in benzoxazinoid accumulation, which were explained by differential benzoxazinoid degradation rather than exudation. Benzoxazinoid exudation modulated microbial diversity in root and rhizospheres during maize growth, but not during subsequent wheat growth, while the chemical fingerprint of benzoxazinoids persisted. Averaged across the field, we did not detect feedbacks on wheat performance and defence, apart from a transient decrease in biomass during vegetative growth. Closer analysis, however, revealed significant feedbacks along the chemical and microbial gradient of the field, with effects gradually changing from negative to positive along the gradient.ConclusionOverall, this study revealed that plant–soil feedbacks differ in strength and direction within a field and that this variation can be explained by standing chemical and microbial gradients. Understanding within‐field soil heterogeneity is crucial for the future exploitation of plant–soil feedbacks in sustainable precision agriculture.

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Introduction: Harnessing positive plant-soil feedbacks via crop rotations is a promising strategy for sustainable agriculture. These feedbacks are often context dependent and how soil heterogeneity explains this variation is unknown. Plants influence soil properties including microbes by exuding specialized metabolites. Benzoxazinoids, specialized metabolites released by cereals such as wheat and maize, can alter rhizosphere microbiota and performance of plants subsequently growing in the exposed soils and are thus an excellent model to study agriculturally relevant plant-soil feedbacks. Materials & methods: To understand local variation in soil properties on benzoxazinoid-mediated plant-soil feedbacks, we conditioned plots with wild-type maize and benzoxazinoid-deficient bx1 mutants in a grid pattern across a field and we then grew winter wheat in the following season. We determined accumulation of benzoxazinoids, root-associated microbial communities, abiotic soil properties and wheat performance in each plot and then assessed their associations. Results: We detected a marked gradient in soil chemical and microbiomes across the field. This gradient resulted in significant differences in benzoxazinoid accumulation, which were explained by differential benzoxazinoid degradation rather than exudation. Benzoxazinoid exudation modulated microbial diversity in root and rhizospheres during maize growth, but not during subsequent wheat growth, while the chemical fingerprint of benzoxazinoids persisted. Averaged across the field, we did not detect feedbacks on wheat performance and defence, apart from a transient decrease in biomass during vegetative growth. Closer analysis however, revealed significant feedbacks along the chemical and microbial gradient of the field, with effects gradually changing from negative to positive along the gradient. Conclusion: Overall, this study revealed that plant-soil feedbacks differ in strength and direction within a field, and that this variation can be explained by standing chemical and microbial gradients. Understanding within-field soil heterogeneity is crucial for the future exploitation of plant-soil feedbacks in sustainable precision agriculture.

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