Plant Biology

Tanacetum vulgare L., known as tansy, is a perennial plant with a highly variable terpenoid composition, with mono- and sesquiterpenoids being the most abundant. The high diversity of terpenoids is known to play an important role in mediating ecological interactions. However, the distribution of terpenoids in different tissues and the inducibility of terpenoids in these tissues by biotic stress are poorly understood. In this study, we investigated the changes in terpenoid profiles and concentrations in different plant organs following treatment of roots with pipecolic acid (Pip). Pipecolic acid is a non-proteinogenic amino acid that triggers defense responses in plants. It is often used to induce systemic resistance (SAR) in plants under controlled conditions. Examination of the tissues showed that the leaves and midribs contained mainly monoterpenoids, while the coarse and fine roots of the plants contained mainly sesquiterpenoids. The rhizomes occupied an intermediate position by presenting the terpenoid profiles of both the midribs and roots but also the unique compounds of its own. Treatment with pipecolic acid led to an increase in the concentration of mono- and sesquiterpenoids in all tissues except rhizomes. However, a significantly higher amount of sesquiterpenoids was formed in root tissues in response to Pip compared to shoots. The metabolic atlas for terpenoids presented here shows that there is an exceptionally strong differentiation of terpenoid patterns and terpenoid contents in the different tissues of tansy. This, together with the differential inducibility by biotic stress, suggests that the chemical diversity of terpenoids may play an important role in the ecological interactions of tansy and in the defense against biotic stressors that feed on the below- and above-ground organs of the plant.

AimIntraspecific variations of specialized metabolites in plants, such as terpenoids, are used to determine chemotypes. Tansy (Tanacetum vulgare L.) exhibits diverse terpenoid profiles, that affect insect communities. However, it is not fully known whether patterns of their chemical composition and associated insects vary on a large scale. Here, we investigated the geographic distribution of mono- and sesquiterpenoid chemotypes in tansy leaves and the effects of these chemotypes on colonization by insect communities across Germany. MethodsWe sampled tansy leaves from 26 sites along a north-south and west-east transect in Germany. Leaves from ten plants with and five plants without aphids was collected from each site. Hexane-extracted metabolites from leaf tissues were analysed by gas chromatography-mass spectrometry (GC-MS). Plant morphological traits, aphid occurrence and abundance, and occurrence of ants were recorded. The effect of plant chemotype, plant morphological parameters, and site parameters such as temperature and precipitation on insect occurrences were analysed. ResultsPlants clustered into four monoterpenoid and four sesquiterpenoid chemotype classes. Monoterpene classes differed in their latitudinal distribution, whereas sesquiterpenes were more evenly distributed across the transect. Aphid and ant occurrence were influenced by monoterpenoids and specific traits. Plants of monoterpenoid class 1 were colonized by Metopeurum fuscoviride and ants significantly more often than expected by chance compared to plants from monoterpenoid class 4. Aphid abundance was negatively affected by host plant height, and increasing average annual temperature positively influenced the occurrence of ants. ConclusionWe found significant geographic differences in the chemodiversity of tansy and show that monoterpenoids affect aphid and ant occurrence, while host plant height can influence aphid abundance. We show that geographic variation in plant chemistry and morphology influences insect communities assemblage on tansy plants.

O_LIPlants can adapt to changing environments by adjusting the production and maintenance of diverse sets of bioactive secondary metabolites. To date, the impact of past climatic conditions relative to other factors such as soil abiotic factors and herbivore pressure on the evolution of plant secondary metabolites is poorly understood, especially for plant roots. C_LIO_LIWe explored associations between root latex secondary metabolites in 63 Taraxacum officinale populations across Switzerland and past climatic conditions, soil abiotic parameters, and root herbivore pressure. To assess the contribution of environmental effects, root secondary metabolites were measured in F0 plants in nature and F2 plants under controlled greenhouse conditions. C_LIO_LIConcentrations of root latex secondary metabolites were most strongly associated with past climatic conditions, while current soil abiotic factors or root herbivore pressure did not show a clear association with root latex chemistry. Results were identical for natural and controlled conditions, suggesting heritable trait variation rather than environmental plasticity as underlying factor. C_LIO_LISynthesis. We conclude that climatic conditions likely play a major role in the evolution of root secondary metabolites. Direct abiotic effects are likely underlying this pattern, hinting at a novel role of root latex metabolites the tolerance of abiotic stress. C_LI

O_LIEuropean beech (Fagus sylvatica L.) is a widely distributed, ecologically and economically important deciduous tree species in European forests, but is increasingly threatened by drought stress. Volatile organic compounds (VOCs) are ubiquitous plant metabolites that may serve as non-invasive biomarkers of drought stress, yet they have rarely been studied in European beech. C_LIO_LIIn this study, we examined VOC responses of European beech to experimental drought across diverse genetic backgrounds in a common garden. The 72 four-year-old beech saplings represented three genetic clusters, seven provenances (geographic seed sources), and 12 maternal seed families. Half of the saplings were assigned to the drought treatment and received no water for 14 days, while the remaining saplings served as controls and were watered as required. VOC profiles, quantified as peak heights of mass spectral features, were measured for all individuals during pre-drought, drought, and rewatering periods. C_LIO_LIWe found that pre-drought VOC profiles, in particular monoterpenes, varied significantly among genetic backgrounds. Experimental drought significantly altered VOC profiles, characterized by increased green leaf volatiles and decreased monoterpenes, oxidized terpenoid derivatives, and other fatty acid derivatives. Reductions in monoterpenes persisted after rewatering, indicating a drought legacy effect. Drought responses were largely conserved across genetic backgrounds, with significant seed family-specific responses detected for only three VOC features. C_LIO_LIOur findings suggest that VOC profiles are genetically structured yet highly plastic under drought and highlight their potential as non-invasive biomarkers for monitoring drought stress in European beech under climate change. C_LI

O_LIThe plant cuticle, multifunctional hydrophobic air-to-plant boundary, vary dramatically among plant species and responds dynamically to environmental and biotic factors. These dynamics, certainly driven by cost-benefit selection, are poorly understood up to now. C_LIO_LI13C labelling and compound-specific isotope analysis open a new avenue to study the cuticle dynamics. We studied side specific leaf cuticular wax regeneration, in different compounds, and separately for intra-(IW) and epicuticular wax (EW), as well as the effect of epicuticular wax removal on the regeneration rate. C_LIO_LIMature leaves that reached the final area before the start of the experiment deposited new IW at first, but quickly equilibrated with EW. Removal of EW accelerated this equilibration but not the net rate of new wax deposition. The n-alkanes had the fastest turnover. Pentacyclic triterpenoids (ursolic acid) had a surprisingly slow turnover due to the large pool but also slow deposition rate. Adaxial and abaxial leaf surfaces exhibited small but consistent differences. C_LIO_LIWe demonstrated for the first time that EW removal does not affect wax deposition rate in evergreen leaf cuticle. We also confirmed that collodion discriminates between EW and IW well and does not inhibit future leaf metabolism and wax deposition in our species. C_LI

A phylogenetic framework explaining plant secondary metabolite diversity is lacking, but metabolite classes could represent adaptations to habitat resource availability. We test the hypothesis that primary adaptive strategies (competitors, C; stress-tolerators, S; ruderals, R) are associated, respectively, with nitrogenous metabolites synthesized in persistent organs (alkaloids), nitrogen-lacking aromatic terpenes and phenolics, and nitrogenous compounds prevalent in reproductive tissues (cyanogenic glucosides and glucosinolates). A matrix was compiled of 1019 species for which secondary metabolite pathways and CSR strategies are known. Accounting for phylogenetic relatedness and native biomes, we found that most phytochemical pathways did not correlate with strategy axes, but certain key associations were evident. C-selection was positively associated with amino acid-derived phenylpropanoids (low phylogenetic relatedness; {lambda} <0.5) and pyrrolizidine alkaloids and galloyl derivatives (high {lambda}), and negatively with N-lacking linear monoterpenes (low {lambda}). Nitrogenous cyanogenic glucosides positively correlated with R-selection (low {lambda}). Terpenoids were widely distributed, but correlated positively with S- and negatively with R-selection (low {lambda}). Twenty-six correlations between phytochemicals and biomes (low {lambda}) were evident. Most secondary metabolite synthesis pathways are widespread, reflecting common roles and obligate defence, and strong phylogenetic effects are often evident. However, the character of phytochemical/adaptive strategy associations agrees with ecological theory and thus reflects adaptation.

Studying natural variation in multi-stress resistance is central for predicting and managing the population dynamics of wild plant species under rapid global change. Yet, it remains a challenging goal in this field to integrate knowledge on the complex biochemical underpinnings for the targeted non-model species. Here, we studied latitudinal divergence in combined drought and heat stress resistance in European populations of the dune plant Cakile maritima, by combining comprehensive plant phenotyping with metabolic profiling via FT-ICR-MS and UPLC-TQ-MS/MS. We observed pronounced constitutive divergence in growth phenology, leaf functional traits and defence chemistry (glucosinolates, alkaloids) among population origins. Most importantly, the magnitude of growth reduction under stress was partly weaker in southern plants and associated with divergence in plastic growth responses (root expansion, leaf abscission) and the stress-induced modulation of primary and specialized metabolites with known central functions not only in plant abiotic but also biotic stress resistance. Our study supports that divergent selection has shaped the constitutive and stress-induced expression of numerous morphological and biochemical functional traits to mediate higher abiotic stress resistance in southern Cakile populations, and highlights that metabolomics is a powerful tool to explore the mechanistic underpinnings of local stress adaptation in non-model species. HighlightPlant defence chemistry and its modulation by abiotic stress exhibits latitudinal clines across natural populations of a coastal plant.

Knowledge of the intraspecific variability of volatiles produced by plants is central for estimating their fluxes from ecosystems and for understanding their evolution in an ecological and phylogenetic context. Past studies suggested that leaf volatile emissions from Cork oak (Quercus suber L.) exhibit a particular high degree of qualitative and quantitative polymorphism. However, the extent of inherent emission variability across its range is not known. We investigated leaf emissions and ecophysiological variables of 241 Cork oak seedlings from ten provenances. To minimize environmental influences, emissions were determined at 30 {degrees}C and saturating light on seed-grown saplings of similar age grown under the same conditions. All individuals, except for three apparent non-emitters, released the same five monoterpenes at a rate of 2559 {+/-} 120 ng m-2 s-1. Northern provenances tended to have higher mean emission rates and lower photosynthetic rates than southern populations, resulting in significant differences in their apparent carbon losses by volatile emissions. Independently, the emission composition varied discontinuously among individuals according to three distinct chemotypes, indicating inherent differences in the activity of two types of monoterpene synthases: one producing -, {beta}-pinene and sabinene, and the other limonene (plus myrcene as a by-product). Chemotype frequencies differed among provenances, particularly between South-Eastern Mediterranean and South-Western Atlantic provenances. Regarding ecophysiological leaf traits, we found no significant difference between chemotypes. The study confirms that Cork oak is a strong monoterpene emitter, showing independent intraspecific variability in emission quality and quantity, with non-emitters being rare. Comparison of the emission variability with those reported for other oak species suggests that an ancestral Pinene/sabine chemotype has diversified within the oak subgenus Cerris during its radiation. This diversification is less pronounced in Cork oak than in other sympatric oaks, possibly due to differential fragmentation and expansion of their ranges in the past.

O_LIThe frequency and length of drought periods are increasing in subtropical and temperate regions worldwide. Epigenetic responses to water stress could be key for plant resilience to this largely unpredictable challenge. Experimental DNA demethylation together with application of a stress factor stands as a suitable strategy to uncover the contribution of epigenetics to plant responses to stress. C_LIO_LIWe analysed leaf cytosine methylation changes in adult plants of the Mediterranean weed, Erodium cicutarium, after seed demethylation with 5-Azacytidine and/or recurrent water stress in a greenhouse. We used bisulfite RADseq (BsRADseq) and a newly reported reference genome for E. cicutarium to characterize methylation changes in a 2x2 factorial design, controlling for plant relatedness. C_LIO_LIIn the long-term, 5-Azacytidine treatment alone caused both hypo and hyper-methylation at individual cytosines, with substantial hypomethylation in CG contexts. In control conditions, drought resulted in a decrease in methylation level in all but CHH contexts. In contrast, the genome of plants that experienced recurrent water stress and had been treated with 5-Azacytidine increased DNA methylation level by ca. 5%. C_LIO_LISeed demethylation and recurrent drought exhibited a highly significant interaction in terms of global and context-specific cytosine methylation supporting an epigenetic contribution in response to stress at molecular level. C_LI

In classic coevolutionary interactions, host plants are thought to accrue novel chemical defenses which are later countered by detoxification strategies and sometimes sequestration in specialist herbivores. We recently discovered that unusual nitrogen- and sulfur-containing (N,S-) cardenolides in some milkweed species are highly toxic, and broken down to less toxic forms which are sequestered by monarch butterflies (Danaus plexippus). Here we isolated and purified five dominant cardenolide toxins from the tropical milkweed, Asclepias curassavica, a globally abundant host plant of monarchs, and fed them to caterpillars individually or in mixture. We hypothesized that the two N,S-cardenolides in A. curassavica (uscharin and voruscharin) would reduce caterpillar growth and sequestration more than other abundant related cardenolides (15-Hydroxy calotropin, frugoside, calactin). Overall, cardenolide treatments caused monarchs to feed more and grow more compared to controls; nonetheless, one N,S-cardenolide (voruscharin) was not stimulatory and caused substantial reductions in growth efficiency. Consuming N,S-cardenolides caused caterpillars to sequester the lowest total amounts of cardenolides, and also reduced their efficiency of sequestration. We next tested the phytochemical diversity hypothesis, that toxin mixtures pose a substantial burden for caterpillars compared to individual compounds provided in equimolar concentrations. We prepared two types of mixtures, one containing equal concentrations of the five compounds and another "realistic mixture" where toxin concentrations reflect their natural proportions in leaves. Mixtures had a negative impact on caterpillar feeding, growth, sequestration, and sequestration efficiency compared to the average of single compounds. The equal and realistic mixtures had similar impacts on feeding and growth, but feeding on the realistic mixture resulted in the lowest sequestration. We conclude that as a result of coevolutionary interactions, even sequestering herbivores may be thwarted by highly specialized plant metabolites such as N,S-cardenolides, and that phytochemical mixtures strengthen plant defense. Toxin mixtures likely challenge detoxification and transport of plant defenses, reducing the herbivores growth and sequestration.

The Mediterranean Basin is a hotspot of plant diversity, with many species producing aromatic essential oils (EOs) that mediate ecological interactions and stress responses. Within Lamiaceae, the genus Thymus shows remarkable chemical variability, yet high intraspecific EO variation often limits its taxonomic resolution. We investigated EO composition, genetic structure, and environmental influences across 39 populations of Thymus sect. Mastichina in the Iberian Peninsula, using GC-MS alongside soil and climatic data to assess drivers of chemical variation and refine taxonomic characterization. We identified 14 major EO compounds, dominated by oxygenated monoterpenes. Most populations exhibited 1,8-cineole-rich chemotypes, yet seven populations showed linalool dominance, and multiple chemotypes often co-occurred within the same populations, revealing high intrapopulation chemical diversity. Minor compounds, including camphor, borneol, and camphene, varied among genetic clusters and were significantly correlated with temperature and precipitation gradients. Differences in EO composition were also detected between ploidy levels and genetic groups, although the major compounds (1,8-cineole and linalool) remained relatively consistent, indicating both conserved and locally adaptive chemical traits. These findings suggest that EO diversity in Thymus sect. Mastichina arises from a complex interplay of environmental conditions, genetic background, and ploidy. Integrating chemical, genetic, and ecological data provides a robust framework for understanding the evolutionary and ecological drivers of EO variation in Mediterranean aromatic plants, with implications for taxonomy, conservation, and the study of adaptive chemical traits.

O_LIBackground and Aims: Grasslands underpin global biodiversity, carbon storage, and ecosystem resilience, yet are increasingly threatened by rising temperatures and water scarcity. Understanding how key physiological traits respond to drought and heat is essential for ensuring grassland function under future conditions. Here, we investigated intraspecific variation in leaf cuticular traits--specifically minimum cuticular conductance after stomatal closure (gmin) and its response to temperature--in six Australian accessions of Themeda triandra spanning a wide climatic gradient. We asked whether gmin and its response to drought reflect climate-of-origin and whether the cuticle shows a thermal threshold (Tp) beyond which conductance rises sharply. C_LIO_LIMethods: Plants from six accessions were grown under well-watered (control) glasshouse conditions and then exposed to drought. We measured gmin in both control and drought treatments and assessed the response of gmin to increasing temperature (30-55 {degrees}C) using fresh fully hydrated leaves (control). Climatic data for each site of origin were used to explore trait-environment relationships. C_LIO_LIKey results: Under well-watered conditions, gmin showed no link to climate-of-origin. Under drought, however, gmin displayed clearer climate-linked patterns: accessions from cooler, wetter regions had lower values, contrary to expectations. Drought responses varied strongly among accessions, ranging from marked reductions to significant increases in gmin. With increasing temperature, gmin declined gradually and no accession exhibited a distinct phase transition, indicating a thermally stable cuticle. C_LIO_LIConclusions: Themeda triandra shows considerable intraspecific diversity in gmin and its response to drought. The clearer alignment between climate of origin and gmin measured under drought conditions suggests drought stress is an important filter for gmin expression. These findings provide a physiological basis for identifying genotypes with enhanced resilience for use in grassland conservation and restoration under a warming, drying climate. C_LI

O_LILeaf minimum conductance (gmin) is important in determining plant responses to drought. However, we do not understand how gmin is related to drought tolerance across species, and how important it is in determining the time to reach critical dehydration levels. C_LIO_LIIn 18 coexisting species from a seasonal tropical forest we quantified gmin to test relationships with early, moderate and severe dehydration thresholds associated with tolerance to turgor loss, breakdown of structural integrity, and disruption of cellular function, respectively. We quantified other functional and hydraulic traits to determine the major axes of trait variation in these species. C_LIO_LIVariation in gmin was the primary determinant of the time to reach critical levels of dehydration, and was unrelated to thresholds for early and severe dehydration. Surprisingly, gmin was negatively related to maximum stomatal conductance, but unrelated to other functional and hydraulic traits. C_LIO_LIThese results highlight the importance of avoiding dehydration via minimizing gmin, and suggest that avoidance, tolerance to early, and severe dehydration represent independent strategies for coping with drought. This would allow coexisting species to balance opportunities for carbon gain with costs of physiological breakdown when faced with varying intensities and durations of drought. C_LI

Environmental changes can trigger phenotypic variation in plants through epigenetic mechanisms, but strong genetic influences on epigenetic variation and phenotypes make it difficult to isolate and study these effects. We investigated phenotypic plasticity using the Lombardy poplar (Populus nigra cv. Italica Duroi), a globaly distributed clonal tree. We surveyed 14 functional traits related to tree growth, ecophysiological and phenological processes in poplar ramets collected along a wide geographical range in Europe and planted under common garden conditions. We investigated whether phenotypic variation was related to geography and historical bioclimatic data of the ramets sites of origin using linear mixed effect models. We found significant differences in among ramets from different geographic origins in tree height, number of stems per ramet and duration of bud flush. However, microenvironmental variation in the common garden, captured via block effects, had an even bigger impact on phenotypic variation than the environmental conditions at the sites of origin. Our results show that phenotypic variation in the ramets might be associated to the climate origin from different climates, suggesting possible epigenetic memory. However, such legacy effects might be quickly outweighed by new environmental conditions.

Based on the hypothesis that variation of the metabolomes of latex is a response to selective pressure and should thus be affected differently from other organs, their variation could provide insight into the defensive chemical selection of plants. Metabolic profiling was utilized to compare tissues of Euphorbia species collected in various regions. The metabolic variation of latexes was much more limited than that of other organs. In all of the species, the levels of polyisoprenes and terpenoids were found to be much higher in latexes than in leaves and roots. Polyisoprenes were also observed to physically delay the contact and growth of pathogens with plant tissues. A secondary barrier composed of terpenes and, in particular, 24-methylenecycloartanol, exhibited antifungal activity. These results, together with the known roles of the enzymes also present in latexes, demonstrate that they are part of a cooperative defense system that comprises both biochemical and physical elements.

To predict and mitigate the effects of climate change on communities and ecosystems, the joint effects of extreme climatic events on species interactions need to be understood. Using the common ragweed (Ambrosia artemisiifolia L.)--leaf beetle (Ophraella communa) system, we investigated the effects of heat wave and elevated CO2 on common ragweed growth, secondary metabolism, and the consequent impacts on the beetle. The results showed that elevated CO2 and heat wave facilitated A. Artemisiifolia growth; further, A. artemisiifolia accumulated large amounts of defensive secondary metabolites. Being fed on A. artemisiifolia grown under elevated CO2 and heat wave conditions resulted in the poor performance of O. communa (high mortality, long development period, and low reproduction). Overall, under elevated CO2, heat wave improved the defensive ability of A. artemisiifolia against herbivores. This implies that heat wave event will relieve harm of A. artemisiifolia to human under elevated CO2. On the other hand, super adaptability to climatic changes may aggravate invasive plant distribution, posing a challenge to the control of invasive plants in the future.

PremiseThe family Annonaceae possesses a broad array of floral phenotypes and pollination specialisations, and are important in the plant-pollinator interactions of tropical rainforests. Although there has been considerable effort to assess their interactions with pollinators, attempts to characterise their visual and olfactory communication channels are scarce. MethodsHere, we investigated the pollination biology of 12 Annonaceae species from five genera, viz. Meiogyne, Monoon, Polyalthia, Pseuduvaria, and Uvaria. Furthermore, their floral colour was characterised by reflectance spectroscopy and floral odour chemistry was assessed using gas chromatography-mass spectrometry. Floral scent was further compared across the whole family using non-metric dimensional scaling plots to identify specialisation in floral odour. ResultsThe Meiogyne species are likely pollinated by small beetles; the Polyalthia and Pseuduvaria species are likely pollinated by beetles and flies; and the Uvaria species is likely pollinated by beetles and bees. Flowers of most species are UV non-reflective, and have various spectral reflectance profile across the remaining visible spectra. Multiple species produce floral odour resembling ripe fruits. The flowers of Meiogyne species and Polyalthia xanthocarpa emitted mostly branched-chain esters, while flowers of Uvaria released mainly straight-chain esters. The Pseuduvaria species instead emitted scent reminiscent of rotten fruits, largely consisting of 2,3-butanediol. The inner petal corrugation in Meiogyne functions as a food reward, and the inner petal growth serves as a nectary gland for Pseuduvaria. ConclusionsOur study identifies the visual and olfactory cues of multiple Annonaceae species and provides insights into how Annonaceae flowers attract different guilds of pollinators.

Heat tolerance determines the vitality of tree species under climate change independently of drought tolerance, but has been much less studied than tree water relations. We studied species-specific differences and the capacity for seasonal heat acclimation in Central Europes naturally most important tree species, Fagus sylvatica, in comparison with two exotic tree species (Fagus orientalis, Pseudotsuga menziesii) that are considered for silvicultural climate change adaptation in managed forests. Foliage of mature trees was incubated at temperatures from 35-50 {degrees}C for up to 4 h to simulate daily heat maxima during heat waves. The maximum quantum yield (Fv/Fm) of photosystem II (PS II) of dark-adapted leaves was measured, because the PS II is particularly sensitive to heat and its functionality can decide on plant survival under heat. Fagus sylvatica was much more tolerant to heat than Pseudotsuga menziesii, but weakly (albeit significantly) less tolerant than Fagus orientalis. Within its limits, Pseudotsuga menziesii showed high seasonal heat acclimation with constantly increasing tolerance during the growing season. Fagus orientalis, but practically not Fagus sylvatica, also acclimated to heat. This makes Fagus orientalis slightly superior over Fagus sylvatica in terms of heat tolerance, whereas the suitability of Pseudotsuga menziesii for silvicultural climate change adaptation is questionable. Strong heat acclimation, but also overall low heat tolerance, in Pseudotsuga menziesii might be the result of evergreenness, which requires the generation of both cold and heat tolerance during the year.

(1) Climate change leads to more extreme weather events. Therefore, a high stress tolerance is becoming more critical for plants, with higher ploidy being reported to lead to higher stress tolerance. Phragmites australis (P. australis) is a target species for paludiculture, i.e. the wet use of peatlands, and well known for its many ploidy levels. (2) We expected octoploid genotypes of P. australis to outperform tetraploid ones in a 15-month mesocosm experiment including a gradient of 0 to 100 days of drought. We used pairs of genotypes differing in ploidy from three different geographic regions. (3) Increasing drought length led to a decrease in growth, biomass, morphological and ecophysiological traits in both ploidy levels, but 4x outperformed 8x in almost all traits under constant water supply (e.g., 2.5-fold more biomass production) and up to moderate drought (about 50 days). Under severe and prolonged drought, both ploidy levels performed equally poorly. (4) Our study suggests that higher ploidy levels do not necessarily outperform lower ploidy levels of P. australis under stressful conditions. In this species, ploidy alone may not explain performance, but the genotype can be as or more important than ploidy.

Davana (Artemisia pallens) is a valuable aromatic herb within the Asteraceae family, highly prized for its essential oil (EO) produced in the aerial parts. However, the root volatiles and their specific composition, and genes responsible for root volatiles have remained unexplored until now. Here, we show that A. pallens roots possess distinct oil bodies and yields [~]0.05% of EO, which is primarily composed of sesquiterpenes {beta}-elemene, neryl isovalerate, {beta}-selinene, and -selinene, and trace amounts of monoterpenes {beta}-myrcene, D-limonene. This shows that, besides aerial parts, roots of davana can also be a source of unique EO. Moreover, we functionally characterized a terpene synthase (ApTPS1) that exhibited high in silico expression in the root transcriptome. The recombinant ApTPS1 showed the formation of {beta}-elemene and germacrene A with E,E-farnesyl diphosphate (FPP) as a substrate. Further, detailed analysis of assay products revealed that {beta}-elemene was the thermal rearrangement product of germacrene A. Furthermore, the functional expression of ApTPS1 in Saccharomyces cerevisiae confirmed the in vivo {beta}-elemene/germacrene A synthase activity of ApTPS1. At the transcript level, ApTPS1 displayed predominant expression in root, with significantly lower level of expression in other tissues. This expression pattern of ApTPS1 positively correlated with the tissue-specific accumulation level of {beta}-elemene. Overall, these findings provide fundamental insights into the EO profile of davana roots, and the contribution of ApTPS1 in the formation of a major root volatile. Main conclusionThe study demonstrated that Artemisia pallens roots can be a source of terpene-rich essential oil and root-specific ApTPS1 forms germacrene-A contributing to major root volatiles.