Crops with weed suppressive root exudates or the direct use of bioherbicidal allelochemicals is a new approach in integrated weed management systems. In this context, the allelopathic activity and chemical composition of root exudates from six genotypes (modern varieties and landraces) of barley were characterized. The phenolic acids appeared to be particularly implicated in the inhibitory action of barley root exudates against Bromus diandrus. The amount of these compounds was higher in sandy substrate than in sandy-clay-loam substrate. Ten phenolic acids and one phenylpropanoid derivative were present, in addition to saponarin, a newly identified flavonoid in barley root exudates. Seven compounds explaining variability in the inhibitory activity of barley roots (stepwise analysis) and one compound detected only in highly allelopathic genotypes were toxic against receiver plants. Most compounds had a greater inhibitory effect on the growth of great brome than the barley genotypes. The synergistic and/or additive effect of the eight compounds appeared to be the source of the toxicity. Benzoic acid, the mixture of compounds, saponarin and salicylic acid were the most efficient compounds against the great brome and the less aggressive against barley. Overall, the results revealed the allelopathic potential of the water-soluble compounds exuded by the roots of living barley plants. These compounds included saponarin, a flavonoid not yet recognized as a barley root allelochemical.

Bacterial volatiles play important roles in mediating beneficial interactions between plants and their associated microbiota. Despite their relevance, bacterial volatiles are mostly studied under laboratory conditions, although these strongly differ from the natural environment bacteria encounter when colonizing plant roots or shoots. In this work, we ask the question whether plant-associated bacteria also emit bioactive volatiles when growing on plant leaves rather than on artificial media. Using four potato-associated Pseudomonas, we demonstrate that potato leaves offer sufficient nutrients for the four strains to grow and emit volatiles, among which 1-undecene and Sulfur compounds have previously demonstrated the ability to inhibit the development of the oomycete Phytophthora infestans, the causative agent of potato late blight. Our results bring the proof of concept that bacterial volatiles with known plant health-promoting properties can be emitted on the surface of leaves and warrant further studies to test the bacterial emission of bioactive volatiles in greenhouse and field-grown plants.

Roots are crucial in plant adaptation through the exudation of various compounds which are influenced and modified by environmental factors. Buckwheat root exudate and root system response to neighbouring plants (buckwheat or redroot pigweed) and how these exudates affect redroot pigweed was investigated. Characterising root exudates in plant–plant interactions presents challenges, therefore a split-root system which enabled the application of differential treatments to parts of a single root system and non-destructive sampling was developed. Non-targeted metabolome profiling revealed that neighbour presence and identity induces systemic changes. Buckwheat and redroot pigweed neighbour presence upregulated 64 and 46 metabolites, respectively, with an overlap of only 7 metabolites. Root morphology analysis showed that, while the presence of redroot pigweed decreased the number of root tips in buckwheat, buckwheat decreased total root length and volume, surface area, number of root tips, and forks of redroot pigweed. Treatment with exudates (from the roots of buckwheat and redroot pigweed closely interacting) on redroot pigweed decreased the total root length and number of forks of redroot pigweed seedlings when compared to controls. These findings provide understanding of how plants modify their root exudate composition in the presence of neighbours and how this impacts each other’s root systems.

Within the plant kingdom, there is an exceptional amount of chemical diversity that has yet to be annotated. It is for this reason that non-targeted analysis is of interest for those working in novel natural products. To increase the number and diversity of compounds observable in root exudate extracts, several workflows which differ at three key stages were compared: 1) sample extraction, 2) chromatography, and 3) data preprocessing. Plants were grown in Hoagland's solution for two weeks, and exudates were initially extracted with water, followed by a 24-h regeneration period with subsequent extraction using methanol. Utilizing the second extraction showed improved results with less ion suppression and reduced retention time shifting compared to the first extraction. A single column method, utilizing a pentafluorophenyl column, paired with high-resolution mass spectrometry ionized and correctly identified 34 mock root exudate compounds, while the dual column method, incorporating a pentafluorophenyl column and a porous graphitic carbon column, retained and identified 43 compounds. In a pooled quality control sample of exudate extracts, the single column method detected 1,444 compounds. While the dual method detected fewer compounds overall (1,050), it revealed a larger number of small polar compounds. Three preprocessing methods (targeted, proprietary, and open source) successfully identified 43, 31, and 38 mock root exudate compounds to confidence level 1, respectively. Enhancing signal strength and analytical method stability involves removing the high ionic strength nutrient solution before sampling root exudate extracts. Despite signal intensity loss, a dual column method enhances compound coverage, particularly for small polar metabolites. Open-source software proves a viable alternative for non-targeted analysis, even surpassing proprietary software in peak picking.

This study investigates the effects of substrate composition on root architecture, plant growth, and allelopathic secondary metabolites, specifically benzoxazinoids (BXs), in the rhizospheres of rye (Secale cereale L.) and redroot pigweed (Amaranthus retroflexus L.). Given the complexities of root exudate analysis, including the influence of substrate on root morphology and exudation, the experiment compared plant growth and BX release in two substrates: glass microbeads and a mixture of clay beads and attapulgite. Rye, pigweed, and co-cultures of the two were grown under controlled conditions, with root and shoot parameters measured to assess substrate suitability. Additionally, UPLC-QTOF-MS was used to analyze BXs in rye and rye–pigweed co-cultures. The results demonstrated that the clay bead and attapulgite mixture provided better growth conditions and was effective for BX extraction, making it a suitable substrate for studying allelopathy in controlled environments. The findings highlight the critical role of substrate composition in both plant development and the study of root exudates, with implications for better understanding of crop–weed interactions and allelopathy.

Root exudates are composed of primary and secondary organic compounds that function as signalling molecules and play important roles in plant-environment interactions. The quantity and composition of root exudates vary depending on the species, genotype, and environmental conditions, including the presence and identity of neighbouring plants. Although cover crops are commonly used in agricultural practices for their ecosystem services, such as weed suppression, pathogen control, and soil structure improvement, studies on their root exudates are limited. Our study provides the first characterization of the root exudates of black oat interacting with weed neighbours, redroot pigweed and blackgrass, as well as with neighbours of the same species, black oat. We investigated how these interactions influence the black oat root exudation patterns. Furthermore, we investigated the impact of black oat presence on neighbours and how exposure to black oat root exudate treatments affects the root traits of weeds. The upregulated compounds detected in root exudates in response to neighbouring plants primarily belonged to the organic oxygen compounds superclass, with most of which identified as amino acids and carbohydrates. In the presence of redroot pigweed, a general increase in root exudation was observed, with amino acids and sugar sulphates being upregulated. The presence of black oat had varying effects among the neighbouring plants. While significant decreases observed and black oat in redroot pigweed root traits, an increase was observed in blackgrass. Similarly, more pronounced effects were observed in redroot pigweed compared to blackgrass upon root exudate application. This study provides a characterization and insights into the dynamic nature of root exudates and their influence on root traits and plant interactions.