Glomus Intraradices Research Articles

The Effects of Rhizophagus irregularis Inoculation on Transcriptome of Medicago lupulina Leaves at Early Vegetative and Flowering Stages of Plant Development.

The study is aimed at revealing the effects of Rhizophagus irregularis inoculation on the transcriptome of Medicago lupulina leaves at the early (second leaf formation) and later (flowering) stages of plant development. A pot experiment was conducted under conditions of low phosphorus (P) level in the substrate. M. lupulina plants were characterized by high mycorrhizal growth response and mycorrhization parameters. Library sequencing was performed on the Illumina HiseqXTen platform. Significant changes in the expression of 4863 (padj < 0.01) genes from 34049 functionally annotated genes were shown by Massive Analysis of cDNA Ends (MACE-Seq). GO enrichment analysis using the Kolmogorov-Smirnov test was performed, and 244 functional GO groups were identified, including genes contributing to the development of effective AM symbiosis. The Mercator online tool was used to assign functional classes of differentially expressed genes (DEGs). The early stage was characterized by the presence of six functional classes that included only upregulated GO groups, such as genes of carbohydrate metabolism, cellular respiration, nutrient uptake, photosynthesis, protein biosynthesis, and solute transport. At the later stage (flowering), the number of stimulated GO groups was reduced to photosynthesis and protein biosynthesis. All DEGs of the GO:0016036 group were downregulated because AM plants had higher resistance to phosphate starvation. For the first time, the upregulation of genes encoding thioredoxin in AM plant leaves was shown. It was supposed to reduce ROS level and thus, consequently, enhance the mechanisms of antioxidant protection in M. lupulina plants under conditions of low phosphorus level. Taken together, the obtained results indicate genes that are the most important for the effective symbiosis with M. lupulina and might be engaged in other plant species.

Analysis of express sequence tags from Arbuscular Mycorrhizal fungi (AMF) express during the pre-symbiotic stage of the fungus

Arbuscular Mycorrhizal Fungi (AMF) are symbiotically associated with the roots of higher plants which enhance the overall nutritional status of plants. It is known that the overall growth and development of plants not only depend upon the performance of specific genotypes but their association is also affected by the various types of microbes. Therefore, the major aim of the study was the isolation and characterization of gene expression during the presymbiotic stage of AMF. In the present study, total RNA was extracted from spores of Rhizophagus irregularis grown under root organ culture (ROC). cDNA library was prepared and expressed sequence tag (EST) analysis was performed. Approximately 150 random clones were selected for EST sequencing analysis. Out of the 150 clones obtained, 1.5% carried inserts of the rRNA gene cluster. After excision, inserts from 72 randomly selected clones were sequenced. Database searches revealed that 74% of the clones had similarities to already known sequences. Putative genes for Ca2+-regulated symbiotic genes, cell cycle regulatory factors and precursors for lipid biosynthesis genes were discovered. A full-length mating gene was discovered using rapid amplification of cDNA ends (RACE). Studies revealed the presence of a putative mating type gene in mycorrhizal species that are evident in the sexual origin of AMF.

Synergistic Interaction of Rhizobium tropici, Rhizophagus irregularis and Serendipita indica in Promoting Snap Bean Growth

The overuse of chemical pesticides and fertilizers in crop farming has led to a decrease in crop quality and negative impacts on soil and the environment. It is crucial to adopt alternative strategies to maintain soil and environmental quality while enhancing crop growth and yield. To explore this, a study was conducted under greenhouse conditions to investigate the effect of Rhizobium tropici CIAT 899 alone, as well as in association with mycorrhizae (Rhizophagus irregularis) and endophytic fungus (Serendipita indica), on the growth, yield, and nutrient status of snap bean plants. At harvest, the rhizobial strain CIAT 899 demonstrated the highest effectiveness. It significantly increased the number of nodules in both Contender and Garrafal Enana varieties by 6.97% and 14.81%, respectively, compared with the control without inoculation. Furthermore, the results indicated that co-inoculation of Rhizobium and symbiotic fungi had positive effects on nitrogen content, phosphorus availability, and overall plant growth. Regardless of the variety, plants inoculated with R. tropici CIAT 899 and Serendipita indica exhibited the highest values for plant growth parameters. This combination resulted in 168% and 135% increases in root dry biomass, as well as 140% and 225% increases in the number of pods for Contender and Garrafal Enana, respectively, compared with the control at harvest. Additionally, this study highlights the potential benefits of combining R. tropici with either Serendipita indica or Rhizophagus irregularis in terms of nitrogen and phosphorus uptake. These symbiotic microorganisms demonstrated synergistic interactions with snap bean plants, leading to improved mineral nutrition and enhanced growth. Overall, these findings suggest that utilizing these symbiotic microorganisms can effectively enhance the mineral nutrition and growth of snap bean plants.

Cooperative interaction of Glomus intraradices with plant growth-promoting rhizobacteria promotes plant development and essential oil yield of Pogostemon cablin and reduces disease occurrence under organic field conditions

Cooperative interaction of Glomus intraradices with plant growth-promoting rhizobacteria promotes plant development and essential oil yield of Pogostemon cablin and reduces disease occurrence under organic field conditions

Can cardiolipins be used as a biomarker for arbuscular mycorrhizal fungi?

Specific biomarker molecules are increasingly being used for detection and quantification in plant and soil samples of arbuscular mycorrhizal (AM) fungi, an important and widespread microbial guild heavily implicated in transfers of nutrients and carbon between plants and soils and in the maintenance of soil physico-chemical properties. Yet, concerns have previously been raised as to the validity of a range of previously used approaches (e.g., microscopy, AM-specific fatty acids, sterols, glomalin-like molecules, ribosomal DNA sequences), justifying further research into novel biomarkers for AM fungal abundance and/or functioning. Here, we focused on complex polar lipids contained in pure biomass of Rhizophagus irregularis and in nonmycorrhizal and mycorrhizal roots of chicory (Cichorium intybus), leek (Allium porrum), and big bluestem (Andropogon gerardii). The lipids were analyzed by shotgun lipidomics using a high-resolution hybrid mass spectrometer. Size range between 1350 and 1550Da was chosen for the detection of potential biomarkers among cardiolipins (1,3-bis(sn-3'-phosphatidyl)-sn-glycerols), a specific class of phospholipids. The analysis revealed a variety of molecular species, including cardiolipins containing one or two polyunsaturated fatty acids with 20 carbon atoms each, i.e., arachidonic and/or eicosapentaenoic acids, some of them apparently specific for the mycorrhizal samples. Although further verification using a greater variety of AM fungal species and samples from various soils/ecosystems/environmental conditions is needed, current results suggest the possibility to identify novel biochemical signatures specific for AM fungi within mycorrhizal roots. Whether they could be used for quantification of both root and soil colonization by the AM fungi merits further scrutiny.

Are Histidine Kinases of Arbuscular Mycorrhizal Fungi Involved in the Response to Ethylene and Cytokinins?

Signals are exchanged at all stages of the arbuscular mycorrhizal (AM) symbiosis between fungi and their host plants. Root-exuded strigolactones are well-known early symbiotic cues, but the role of other phytohormones as interkingdom signals has seldom been investigated. Here we focus on ethylene and cytokinins, for which candidate receptors have been identified in the genome of the AM fungus Rhizophagus irregularis. Ethylene is known from the literature to affect asymbiotic development of AM fungi, and in the present study, we found that three cytokinin forms could stimulate spore germination in R. irregularis. Heterologous complementation of a Saccharomyces cerevisiae mutant strain with the candidate ethylene receptor RiHHK6 suggested that this protein can sense and transduce an ethylene signal. Accordingly, its N-terminal domain expressed in Pichia pastoris displayed saturable binding to radiolabeled ethylene. Thus, RiHHK6 displays the expected characteristics of an ethylene receptor. In contrast, the candidate cytokinin receptor RiHHK7 did not complement the S. cerevisiae mutant strain or Medicago truncatula cytokinin receptor mutants and seemed unable to bind cytokinins, suggesting that another receptor is involved in the perception of these phytohormones. Taken together, our results support the hypothesis that AM fungi respond to a range of phytohormones and that these compounds bear multiple functions in the rhizosphere beyond their known roles as internal plant developmental regulators. Our analysis of two phytohormone receptor candidates also sheds new light on the possible perception mechanisms in AM fungi. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Effects of inoculation with four mycorrhizal species on seed phenolic and fatty acids of sesame plants grown under different irrigation regimes

This study evaluated the interaction effects of irrigation level (well-watered and water stress conditions) and inoculation by different mycorrhizal species (non-inoculated, Funneliformis mosseae, Rhizophagus irregularis, Claroideoglomus claroideum, and Glomus fasciculatum) on mycorrhizal colonization, antioxidant activity, seed yield and oil quality of two sesame cultivars (Yekta and Naz). Water deficit decreased mycorrhizal colonization, seed yield and oil concentration but increased antioxidant activity and seed total phenol and flavonoid concentrations. However, mycorrhizal inoculation increased antioxidant activity, seed yield, oil concentration and total phenolic and flavonoids. The lowest reduction by water stress and the highest increase by inoculation in seed yield were observed in Naz plants inoculated by Cl. claroideum. Principal component analysis showed the highest differentiation effect of water stress compared to mycorrhizal inoculation on both cultivars, indicating the relative sensitivity of the two cultivars to water deficit. However, the application of different species of mycorrhizal fungi versus the non-inoculation conditions was somewhat discriminative. In terms of fatty acids, in most cases, water stress increased oleic, palmitic and stearic acids and decreased linoleic and linolenic acids but inoculation increased oleic and linoleic acids and decreased linolenic, palmitic and stearic acids. Regarding phenolic and flavonoids components, the contents of chlorogenic and caffeic acids were increased by water stress but no consistent trend was noted in response to water stress for the other compounds. Mycorrhizal inoculation generally decreased chlorogenic acid but increased gallic, caffeic, p-coumaric, and ferulic acids. In conclusion, the results of the present study may help to increase the level of valuable compounds in sesame for further pharmaceutical purposes under water stress conditions and mycorrhizal symbiosis.

Synergism of microorganisms and seaweed extract on vegetative growth, yield and quality of cucumber fruit

Natural biostimulants, such as microorganisms and seaweed extracts, are used in agriculture to improve crop yields with a sustainable approach. However, the interaction effects between different biostimulants have not been thoroughly investigated. The objective of this work was to evaluate the synergistic effects of microorganisms (Glomus intraradices and Azospirillum brasilense) and seaweed extract (Ascophyllum nodosum and Macrocytis pyrifera) on growth, yield and fruit quality of cucumber plants under soilless cultivation. Two doses of microorganisms (0 and 10 spores mL-1 + 106 CFU) and four concentrations of seaweed extract (0, 250, 500 and 2500 ppm) were evaluated. The experimental design used was a randomized complete block design with a factorial arrangement (2 x 4), with four replications per treatment. The results showed that the equatorial diameter of the fruit, yield and titratable acidity improve when applying microorganisms and seaweed extract in combination; however, when applying the two biostimulants the values ​​of plant height, stem diameter, number of leaves, root length, biomass (fresh and dry), firmness, total soluble solids, vitamin C, chlorophyll (a and total) and indices of color (L* and b*) did not exceed those obtained when the biostimulants were applied individually. The combined application of microorganisms and seaweed extract improves cucumber yield, but not vegetative growth and, except for titratable acidity, fruit quality attributes.

Integrative application of biochar and arbuscular mycorrhizal fungi for enhanced chromium resistance in Medicago sativa

Soil chromium (Cr) contamination has become an environmental problem of global concern. However, the joint effects of combined utilization of biochar and arbuscular mycorrhizal (AM) fungal inoculum, which are considered as two promising remediation strategies of soil heavy metal pollutions, on plant Cr resistance are still poorly understood. In this study, a two-factor pot experiment was conducted to investigate how biochar and AM fungus Rhizophagus irregularis regulate Medicago sativa growth, physiological trait, nutrient and Cr uptake, relevant gene expressions, soil properties, and Cr speciation, independently or synergistically. The results showed that biochar notably decreased AM colonization, while biochar and AM fungus could simultaneously increase plant dry biomass. The greatest growth promotion was observed in mycorrhizal shoots at the highest biochar level (50 g kg−1 soil) by 91 times. Both biochar application and AM fungal inoculation enhanced plant photosynthesis and P nutrition, but the promoting effects of AM fungus on them were significantly greater than that of biochar. In addition, the combined application of biochar and AM fungus dramatically reduced shoot and root Cr concentrations by up to 92 % and 78 %, respectively, compared to the non-amended treatment. Meanwhile, down-regulated expressions were observed for metal chelating-related genes. Furthermore, Cr translocation from roots to shoots was reduced by both two soil amendments. Transcriptional levels of genes involved in reactive oxygen species and proline metabolisms were also regulated by biochar application and AM fungal colonization, leading to alleviation of Cr phytotoxicity. Furthermore, AM fungal inoculation slightly elevated soil pH but decreased plant-available soil P, which was, by contrast, lifted by biochar addition. The combined application reduced soil acid-extractable Cr concentration by 40 %. This study provides new insights into comprehensively understanding of the mechanisms of biochar and AM fungi combination on improving plant Cr tolerance.

Effect of Inoculation with Rhizophagus irregularis on common bean (Phaseolus vulgaris L) under phosphorus level application in South-Kivu, Eastern DRC

The positive effects of arbuscular mycorrhizal fungi (AMF) on yield and phosphorus uptake have already been widely studied. However, the response of common bean (Phaseolus vulgaris L) to mycorrhizal inoculation under phosphorus supply conditions is still poorly documented in South-Kivu where the use of fungal biofertilizers is not yet tested. This study was initiated to determine the effect of mycorrhizal inoculation and increasing phosphorus doses on common bean performance and yield in South Kivu in order to reduce phosphate input. The study was conducted during the A 2021 cropping season at the experimental field of the “Université Evangélique en Afriqu” in Kashusha, Kabare territory. A split plot design was used to compare two levels of mycorrhizal inoculation (with inoculation and without inoculation) and three rates of increasing phosphorus (0, 30, 60 and 120 kg P ha-1). Growth, yield and mycorrhizal colonization parameters were recorded. Inoculation with R irregularis significantly improved root mycorrhization rate, biomass, yield and harvest index of bean at 0 Kg P ha-1 and 30 Kg P ha-1. The performance of bean plants inoculated with R irregularis, in terms of collar diameter, above-ground biomass, total biomass, yield and harvest index at 30 kg P ha-1, was superior to that non-inoculated and inoculated plants at the doses of 60 kg P ha-1 and 120 kg P ha-1, suggesting the potential of AMF in reducing phosphate fertilizer input. The results indicate that P levels significantly affected the mycorrhization rate of the bean plants. The application of 60 and 120 kg P ha-1 drastically reduced the mycorrhization rate of bean confirming the influence of inorganic P on the establishment of mycorrhizal symbiosis. Therefore, the responses evoked by AMF were also dependent on the applied P dose. Mycorrhizal inoculation with R irregularis could be an important lever to boost bean yield and ensure phosphate fertilizer saving in ferralitic soils of Kashusha.

Monoterpene glucosides in Eustoma grandiflorum roots promote hyphal branching in arbuscular mycorrhizal fungi.

Host plant-derived strigolactones trigger hyphal branching in arbuscular mycorrhizal (AM) fungi, initiating a symbiotic interaction between land plants and AM fungi. However, our previous studies revealed that gibberellin-treated lisianthus (Eustoma grandiflorum, Gentianaceae) activates rhizospheric hyphal branching in AM fungi using unidentified molecules other than strigolactones. In this study, we analyzed independent transcriptomic data of E. grandiflorum and found that the biosynthesis of gentiopicroside (GPS) and swertiamarin (SWM), characteristic monoterpene glucosides in Gentianaceae, was upregulated in gibberellin-treated E. grandiflorum roots. Moreover, these metabolites considerably promoted hyphal branching in the Glomeraceae AM fungi Rhizophagus irregularis and Rhizophagus clarus. GPS treatment also enhanced R. irregularis colonization of the monocotyledonous crop chive (Allium schoenoprasum). Interestingly, these metabolites did not provoke the germination of the root parasitic plant common broomrape (Orobanche minor). Altogether, our study unveiled the role of GPS and SWM in activating the symbiotic relationship between AM fungi and E. grandiflorum.

Enhanced production of select phytocannabinoids in medical Cannabis cultivars using microbial consortia.

The root microbiome of medical cannabis plants has been largely unexplored due to past legal restrictions in many countries. Microbes that live on and within the tissue of Cannabis sativa L. similar to other plants, provide advantages such as stimulating plant growth, helping it absorb minerals, providing protection against pathogen attacks, and influencing the production of secondary metabolites. To gain insight into the microbial communities of C. sativa cultivars with different tetrahydrocannabinol (THC) and cannabidiol (CBD) profiles, a greenhouse trial was carried out with and without inoculants added to the growth substrate. Illumina MiSeq metabarcoding was used to analyze the root and rhizosphere microbiomes of the five cultivars. Plant biomass production showed higher levels in three of five cultivars inoculated with the arbuscular mycorrhizal fungus Rhizophagus irregularis and microbial suspension. The blossom dry weight of the cultivar THE was greater when inoculated with R. irregularis and microbial suspension than with no inoculation. Increasing plant biomass and blossom dry weight are two important parameters for producing cannabis for medical applications. In mature Cannabis, 12 phytocannabinoid compounds varied among cultivars and were affected by inoculants. Significant differences (p ≤ 0.01) in concentrations of cannabidivarinic acid (CBDVA), cannabidivarin (CBDV), cannabigerol (CBG), cannabidiol (CBD), and cannabigerolic acid (CBGA) were observed in all Cannabis cultivars when amended with F, K1, and K2 inoculants. We found microbes that were shared among cultivars. For example, Terrimicrobium sp., Actinoplanes sp., and Trichoderma reesei were shared by the cultivars ECC-EUS-THE, CCL-ECC, and EUS-THE, respectively. Actinoplanes sp. is a known species that produces phosphatase enzymes, while Trichoderma reesei is a fungal train that produces cellulase and contributes to organic matter mineralization. However, the role of Terrimicrobium sp. as an anaerobic bacterium remains unknown. This study demonstrated that the use of inoculants had an impact on the production of phytocannabinoids in five Cannabis cultivars. These inoculants could have useful applications for optimizing cannabis cultivation practices and increasing the production of phytocannabinoids.

Seed Coating of Native Rhizophagus irregularis and Funneliformis sp. for Improved Production and Moisture Stress Mitigation in Finger Millet, Eleusine coracana

Seed Coating of Native Rhizophagus irregularis and Funneliformis sp. for Improved Production and Moisture Stress Mitigation in Finger Millet, Eleusine coracana

Arbuscular mycorrhiza fungi Rhizophagus irregularis regulate iron and zinc homeostasis in tobacco by mediating the expression of Yellow Stripe-Like 7

Arbuscular mycorrhizal (AM) participate in fungi-plant symbiotic relationships and mediate the acquisition of soil-borne nutrients by host plants. However, little is known about how AM colonization affects the homeostasis of iron (Fe) and zinc (Zn) in the host plant. Here, we assessed the performance of mycorrhizal growth, oxidative defense, nutrient accumulation, and key gene responses in AM-colonized tobacco (Nicotiana tabacum L.) under different nutrient stress. The total mycorrhizal frequency of AM fungi on tobacco roots reached 86 % and 75 % under low and high phosphorus (P) treatments, respectively. Under Fe/Zn stress, AM alleviated the inhibition of tobacco biomass accumulation, decreased the content of lipid peroxides, and upregulated enzymatic antioxidant capability. In AM-colonized tobacco, Fe/Zn accumulation was increased under deficient conditions and decreased under surplus conditions. Tobacco uptake of P, Fe, and Zn was impaired by excess nutrient concentrations and improved at low concentrations, and this phenomenon was supported by AM colonization. Fe/Zn deficiency upregulated genes related to Fe/Zn uptake, and Fe/Zn excess upregulated genes related to heavy metal efflux, with AM colonization further promoting these upregulations. Notely, we found a possible crosstalk node, Yellow Stripe-Like 7 (NtYSL7), in the AM and Fe/Zn homeostasis regulatory network. NtYSL7 has a plasma membrane-localized oligopeptide transporter domain. NtYSL7 is strongly induced by AM colonization, deficiency of Fe, and surplus Zn, and interacts with many Fe/Zn transporters. AM assists host plants in maintaining Fe/Zn homeostasis by improving nutrient accumulation, redox homeostasis, and transport mechanisms, and NtYSL7 is likely important within the crosstalk network of AM, Fe, and Zn nutrient homeostasis.

Regulation of mycorrhizal colonization under stress in tomato depends on symbiotic efficiency

The mutualistic symbiosis between plants and arbuscular mycorrhizal (AM) fungi is based on a balanced nutrient exchange between both partners, with the plant achieving improved nutrition and stress tolerance. The symbiosis is finely-tuned according to plant’s needs and surrounding conditions, usually through phytohormonal signaling. Thus, environmental conditions or stress factors modulating phytohormone signaling may influence the symbiosis. This study compares the colonization abilities of 2 AM fungal species, Funneliformis mosseae and Rhizophagus irregularis, independently or in combination, in tomato plants subjected to different stress conditions. These included salt stress and systemic defense activation by aboveground application of the defense-related hormones methyl jasmonate, abscisic acid and salicylic acid. The results show that root colonization by the two fungal species differs depending on the stress treatment. Nutrient and transcriptional analyses revealed that changes in colonization correlated with differential regulation of nutrient exchange, plant defensive responses, and symbiosis regulatory genes. Specifically, under salt stress R. irregularis colonization decreased, while F. mosseae colonization was promoted. These differential regulation of colonization under stress positively correlated with changes in the functionality of the symbiosis. Overall, the results support that the benefits provided by each AM fungi influence carbon reward and determines the control of root colonization by the host plant.

Mycorrhizal response of Solanum tuberosum to homokaryotic versus dikaryotic arbuscular mycorrhizal fungi.

Arbuscular mycorrhizal fungi (AMF) are obligate plant symbionts of most land plants. In these organisms, thousands of nuclei that are either genetically similar (homokaryotic) or derived from two distinct parents (dikaryotic) co-exist in a large syncytium. Here, we investigated the impact of these two nuclear organizations on the mycorrhizal response of potatoes (Solanum tuberosum) by inoculating four potato cultivars with eight Rhizophagus irregularis strains individually (four homokaryotic and four dikaryotic). By evaluating plant and fungal fitness-related traits four months post inoculation, we found that AMF genetic organization significantly affects the mycorrhizal response of host plants. Specifically, homokaryotic strains lead to higher total, shoot, and tuber biomass and a higher number of tubers, compared to dikaryotic strains. However, fungal fitness-related traits showed no clear differences between homokaryotic and dikaryotic strains. Nucleotype content analysis of single spores confirmed that the nucleotype ratio of AMF heterokaryon spores can shift depending on host identity. Together, these findings continue to highlight significant ecological differences derived from the two distinct genetic organizations in AMF.

Comparative RNA sequencing-based transcriptome profiling of ten grapevine rootstocks: shared and specific sets of genes respond to mycorrhizal symbiosis.

Arbuscular mycorrhizal symbiosis improves water and nutrient uptake by plants and provides them other ecosystem services. Grapevine is one of the major crops in the world. Vitis vinifera scions generally are grafted onto a variety of rootstocks that confer different levels of resistance against different pests, tolerance to environmental stress, and influence the physiology of the scions. Arbuscular mycorrhizal fungi are involved in the root architecture and in the immune response to soil-borne pathogens. However, the fine-tuned regulation and the transcriptomic plasticity of rootstocks in response to mycorrhization are still unknown. We compared the responses of 10 different grapevine rootstocks to arbuscular mycorrhizal symbiosis (AMS) formed with Rhizophagus irregularis DAOM197198 using RNA sequencing-based transcriptome profiling. We have highlighted a few shared regulation mechanisms, but also specific rootstock responses to R. irregularis colonization. A set of 353 genes was regulated by AMS in all ten rootstocks. We also compared the expression level of this set of genes to more than 2000 transcriptome profiles from various grapevine varieties and tissues to identify a class of transcripts related to mycorrhizal associations in these 10 rootstocks. Then, we compared the response of the 351 genes upregulated by mycorrhiza in grapevine to their Medicago truncatula homologs in response to mycorrhizal colonization based on available transcriptomic studies. More than 97% of the 351M. truncatula-homologous grapevine genes were expressed in at least one mycorrhizal transcriptomic study, and 64% in every single RNAseq dataset. At the intra-specific level, we described, for the first time, shared and specific grapevine rootstock genes in response to R. irregularis symbiosis. At the inter-specific level, we defined a shared subset of mycorrhiza-responsive genes.

Arbuscular mycorrhizal fungi impact the production of alkannin/shikonin and their derivatives in Alkanna tinctoria Tausch. grown in semi-hydroponic and pot cultivation systems.

Alkanna tinctoria Tausch. is a medicinal plant well-known to produce important therapeutic compounds, such as alkannin/shikonin and their derivatives (A/Sd). It associates with arbuscular mycorrhizal fungi (AMF), which are known, amongst others beneficial effects, to modulate the plant secondary metabolites (SMs) biosynthesis. However, to the best of our knowledge, no study on the effects of AMF strains on the growth and production of A/Sd in A. tinctoria has been reported in the literature. Here, three experiments were conducted. In Experiment 1, plants were associated with the GINCO strain Rhizophagus irregularis MUCL 41833 and, in Experiment 2, with two strains of GINCO (R. irregularis MUCL 41833 and Rhizophagus aggregatus MUCL 49408) and two native strains isolated from wild growing A. tinctoria (R. irregularis and Septoglomus viscosum) and were grown in a semi-hydroponic (S-H) cultivation system. Plants were harvested after 9 and 37 days in Experiment 1 and 9 days in Experiment 2. In Experiment 3, plants were associated with the two native AMF strains and with R. irregularis MUCL 41833 and were grown for 85 days in pots under greenhouse conditions. Quantification and identification of A/Sd were performed by HPLC-PDA and by HPLC-HRMS/MS, respectively. LePGT1, LePGT2, and GHQH genes involved in the A/Sd biosynthesis were analyzed through RT-qPCR. In Experiment 1, no significant differences were noticed in the production of A/Sd. Conversely, in Experiments 2 and 3, plants associated with the native AMF R. irregularis had the highest content of total A/Sd expressed as shikonin equivalent. In Experiment 1, a significantly higher relative expression of both LePGT1 and LePGT2 was observed in plants inoculated with R. irregularis MUCL 41833 compared with control plants after 37 days in the S-H cultivation system. Similarly, a significantly higher relative expression of LePGT2 in plants inoculated with R. irregularis MUCL 41833 was noticed after 9 versus 37 days in the S-H cultivation system. In Experiment 2, a significant lower relative expression of LePGT2 was observed in native AMF R. irregularis inoculated plants compared to the control. Overall, our study showed that the native R. irregularis strain increased A/Sd production in A. tinctoria regardless of the growing system used, further suggesting that the inoculation of native/best performing AMF is a promising method to improve the production of important SMs.

Non-targeted metabolomics analysis reveals the mechanism of arbuscular mycorrhizal symbiosis regulating the cold-resistance of Elymus nutans.

Elymus nutans is a perennial grass of the Gramineae family. Due to its cold-resistance and nutrition deficiency tolerance, it has been applied to the ecological restoration of degraded alpine grassland on the Qinghai-Tibet Plateau. As an important symbiotic microorganism, arbuscular mycorrhizal fungi (AMF) have been proven to have great potential in promoting the growth and stress resistance of Gramineae grasses. However, the response mechanism of the AMF needs to be clarified. Therefore, in this study, Rhizophagus irregularis was used to explore the mechanism regulating cold resistance of E. nutans. Based on pot experiments and metabolomics, the effects of R. irregularis were investigated on the activities of antioxidant enzyme and metabolites in the roots of E. nutans under cold stress (15/10°C, 16/8 h, day/night). The results showed that lipids and lipid molecules are the highest proportion of metabolites, accounting for 14.26% of the total metabolites. The inoculation with R. irregularis had no significant effects on the activities of antioxidant enzyme in the roots of E. nutans at room temperature. However, it can significantly change the levels of some lipids and other metabolites in the roots. Under cold stress, the antioxidant enzyme activities and the levels of some metabolites in the roots of E. nutans were significantly changed. Meanwhile, most of these metabolites were enriched in the pathways related to plant metabolism. According to the correlation analysis, the activities of antioxidant enzyme were closely related to the levels of some metabolites, such as flavonoids and lipids. In conclusion, AMF may regulate the cold-resistance of Gramineae grasses by affecting plant metabolism, antioxidant enzyme activities and antioxidant-related metabolites like flavonoids and lipids. These results can provide some basis for studying the molecular mechanism of AMF regulating stress resistance of Gramineae grasses.

Phytomanagement of Zn- and Cd-Contaminated Soil: Helianthus annuus Biomass Production and Metal Remediation Abilities with Plant-Growth-Promoting Microbiota Assistance

Mining and industrial activity are contributing to the increase in heavy metal (HM) pollution in soils. Phytoremediation coupled to selected rhizosphere microbiota is an environmentally friendly technology designed to promote HM bioremediation in soils. In this study, sunflower (Helianthus annuus L.) was used together with Rhizophagus irregularis, an arbuscular mycorrhizal fungi (AMF), and Cupriavidus sp. strain 1C2, a plant growth promoting rhizobacteria (PGPR), as a phytoremediation strategy to remove Zn and Cd from an industrial soil (599 mg Zn kg−1 and 1.2 mg Cd kg−1). The work aimed to understand if it is possible to gradually remediate the tested soil while simultaneously obtaining significant yields of biomass with further energetic values by comparison to the conventional growth of the plant in agricultural (non-contaminated) soil. The H. annuus biomass harvested in the contaminated industrial soil was 17% lower than that grown in the agricultural soil—corresponding to yields of 19, 620, 199 and 52 g m−2 of roots, stems, flowers and seeds. It was possible to remove ca. 0.04 and 0.91% of the Zn and Cd of the industrial soil, respectively, via the HM accumulation on the biomass produced. The survival of applied microbiota was indicated by a high root colonization rate of AMF (about 50% more than in non-inoculated agricultural soil) and identification of strain 1C2 in the rhizosphere at the end of the phytoremediation assay. In this study, a phytoremediation strategy encompassing the application of an energetic crop inoculated with known beneficial microbiota applied to a real contaminated soil was successfully tested, with the production of plant biomass with the potential for upstream energetic valorisation purposes.