Seedling ectomycorrhization is central to conifer forest restoration: a case study from Kashmir Himalaya.

Over the past few decades, many countries have attempted to carry out forest landscape restoration over millions of hectares of degraded land. Such efforts, however, have met with limited success because of several factors, including a lack of adequate emphasis on ectomycorrhization of the nursery seedlings. A similar scenario is seen in the Kashmir Himalaya, where the natural regeneration of degraded forests is poor despite ample restoration efforts by forest managers. To overcome this challenge, we identified two promising ectomycorrhizal species, namely Clitocybe nuda and Cortinarius distans, for their use in ectomycorrhization of seedlings of three common conifers, namely Abies pindrow, Cedrus deodara, and Picea smithiana. Laboratory studies were carried out to investigate the requirements for optimum mycelial growth of these ectomycorrhizal fungi. Best ECM mycelial growth was obtained in the basic MMN medium containing glucose as the source of carbon and nitrogen in ammonium form. C. distans showed higher growth than C. nuda across all the treatments and also proved significantly more effective in enhancing the survival and growth of the conifer host plant seedlings. The present study resulted in standardizing the requirements for mass inoculum production of the two mycobionts which could help in successful forest restoration programmes.

Mycorrhizal symbiosis promotes the nutrient content accumulation and affects the root exudates in maize.

BACKGROUND: Arbuscular mycorrhizal fungi (AMF) are a group of important symbiotic microorganisms found in ecosystems. Maize is the second most produced food crop globally. To investigate the mechanisms by which mycorrhizal symbiosis improves maize yields, the effects of mycorrhizal symbiosis on root vigor, nutrient accumulation in various tissues, and root exudates were investigated. We propose the following hypothesis: The secretion of organic acids in root exudates has antagonistic or synergistic effects, which are related to the rhizosphere environment. AMF symbiosis will enhance this effect. RESULT: Rhizophagus aggreatus, Claroideoglomus etunicatum, and Funneliformis mosseae were used to inoculate maize plants separately; meanwhile, maize was inoculated with the above three fungi together for another processing. The plant tissues were sampled at five growth stages: V12 (twelve-leaf), VT (Tassel), R1 (Silking), R2 (Blister), and R4 (Dough stage). The root vigor, and nutrient content in different maize organs and organic acids in root exudates were determined in these stages. The results show that mycorrhizal symbiosis significantly improved the root vigor of maize, especially for plants inoculated with F. mosseae. AMF symbiosis significantly increased N, P, and K accumulation. Mixed inoculation with arbuscular mycorrhizal fungi significantly promoted the accumulation of N and K in maize. P accumulation was significantly promoted by C. etunicatum inoculation. Mycorrhizal symbiosis reduced the levels of protocatechuic, vanillic, citric, and ferulic acid in maize root exudates and increased the levels of p-hydroxybenzoic and caffeic acid. Except for syringic, chlorogenic and succinic acid, the levels of other organic acids in root exudates were higher in plants inoculated with F. mosseae than in other treatments. CONCLUSION: This study demonstrates that mycorrhizal symbiosis improves root vigor and promotes nutrient accumulation at various sites; in addition, mycorrhizal symbiosis affects the content of organic acids in root exudates.

Experimental validation under controlled conditions of real time PCR to quantify arbuscular mycorrhizal colonization in root.

Mycorrhizal colonization of roots is traditionally evaluated by empirical methods, such as root microscopy. We compared this method with data from using a real time PCR technique, and determined the correlation between methods, indicating particularities of a promising system for a quick and accurate molecular diagnostic of arbuscular mycorrhization.

Microbial consortium inoculant increases pasture grasses yield in low-phosphorus soil by influencing root morphology, rhizosphere carboxylate exudation and mycorrhizal colonisation.

BACKGROUND: Pasture farming in south-western Australia is challenged by nutrient-poor soils. We assessed the impact of microbial consortium inoculant (MI) and rock mineral fertiliser (MF) on growth, nutrient uptake, root morphology, rhizosphere carboxylate exudation and mycorrhizal colonisation in three pasture grasses - tall fescue (Festuca arundinacea L.), veldt grass (Ehrharta calycina Sm.) and tall wheatgrass (Thinopyrum ponticum L.) grown in low-phosphorus (P) sandy soil in a glasshouse for 30 and 60 days after sowing (DAS). RESULTS: Veldt grass produced the highest specific root length and smallest average root diameter in both growth periods, and had similar shoot weight, root surface area and fine root length (except at 30 DAS) to tall fescue. Compared with the control, MI alone or combined with MF significantly increased shoot and root biomass (except root biomass at 30 DAS), likely due to the significant increases in root surface area and fine root length. Plants supplied with MI + MF had higher shoot N and P contents than those in the MI and the control treatments at 60 DAS. Malate, citrate and trans-aconitate were the major rhizosphere carboxylates exuded at both 30 and 60 DAS. Malate exudation varied among species and treatments in both growth periods, but citrate exudation was consistently higher in the low-P treatments (control and MI) than the MF and MI + MF treatments. CONCLUSION: Microbial consortium inoculant can positively influence pasture production in low-P soil by increasing root surface area and fine root length, whereas exudation of nutrient-mobilising carboxylates (citrate) is dependent more on soil P supply than microbial consortium inoculant. © 2021 Society of Chemical Industry.

Effects of arbuscular mycorrhizal inoculation on the growth, photosynthesis and antioxidant enzymatic activity of Euonymus maackii Rupr. under gradient water deficit levels.

The role of arbuscular mycorrhizal (AM) fungus (Rhizophagus intraradices) in the amelioration of the water deficit-mediated negative influence on the growth, photosynthesis, and antioxidant system in Euonymus maackii Rupr. was examined. E. maackii seedlings were subjected to 5 water deficit levels, soil water contents of 20%, 40%, 60%, 80% and 100% field capacity (FC), and 2 inoculation treatments, with and without AM inoculation. The water deficit increasingly limited the seedling height, biomass accumulation in shoots and roots, chlorophyll content, gas exchange and chlorophyll fluorescence parameters with an increasing water deficit level. In addition, water deficit stimulated the activities of antioxidant enzymes, including superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT), in both shoots and roots, except under 20% FC conditions. E. maackii seedlings under all water deficit conditions formed symbiosis well with AM fungi, which significantly ameliorated the drought-mediated negative effect, especially under 40% and 60% FC conditions. Under 40% to 80% FC conditions, AM formation improved seedling growth and photosynthesis by significantly enhancing the biomass accumulation, chlorophyll content and assimilation. Mycorrhizal seedlings showed better tolerance and less sensitivity to a water deficit, reflected in the lower SOD activities of shoots and roots and CAT activity of shoots under 40% and 60% FC conditions. Downregulation of the antioxidant system in mycorrhizal seedlings suggested better maintenance of redox homeostasis and protection of metabolism, including biomass accumulation and assimilation. All the results advocated the positive role of R. intraradices inoculation in E. maackii against a water deficit, especially under 40% FC, which suggested the distinct AM performance in drought tolerance and the potential role of the combination of E. maackii-AM fungi in ecological restoration in arid regions.

Combined inoculation with dark septate endophytes and arbuscular mycorrhizal fungi: synergistic or competitive growth effects on maize?

BACKGROUND: Effects on maize were assessed of dual inoculation with arbuscular mycorrhizal fungi (AMF) and dark septate endophytes (DSE) isolated from other plant species. METHODS: Suspensions of DSE isolated from Stipa krylovii were prepared at different densities (2, 4, and 8 × 105 CFU mL- 1) and inoculated separately (AMF or DSE) or together (AMF + DSE), to explore their effects on maize growth. RESULTS: Inoculation with AMF or medium and high densities of DSE and combined inoculation (AMF + DSE) increased plant above-ground growth and altered root morphology. Differences in plant growth were attributable to differences in DSE density, with negative DSE inoculation responsiveness at low density. AMF promoted plant above-ground growth more than DSE and the high density of DSE promoted root development more than AMF. Combined inoculation might lead to synergistic growth effects on maize at low density of DSE and competitive effects at medium and high DSE densities. CONCLUSIONS: AMF and DSE co-colonized maize roots and they had positive effects on the host plants depending on DSE density. These findings indicate the optimum maize growth-promoting combination of AMF and DSE density and provide a foundation for further exploration of potentially synergistic mechanisms between AMF and DSE in physiological and ecological effects on host plants.

MycoRed: Betalain pigments enable in vivo real-time visualisation of arbuscular mycorrhizal colonisation.

Arbuscular mycorrhiza (AM) are mutualistic interactions formed between soil fungi and plant roots. AM symbiosis is a fundamental and widespread trait in plants with the potential to sustainably enhance future crop yields. However, improving AM fungal association in crop species requires a fundamental understanding of host colonisation dynamics across varying agronomic and ecological contexts. To this end, we demonstrate the use of betalain pigments as in vivo visual markers for the occurrence and distribution of AM fungal colonisation by Rhizophagus irregularis in Medicago truncatula and Nicotiana benthamiana roots. Using established and novel AM-responsive promoters, we assembled multigene reporter constructs that enable the AM-controlled expression of the core betalain synthesis genes. We show that betalain colouration is specifically induced in root tissues and cells where fungal colonisation has occurred. In a rhizotron setup, we also demonstrate that betalain staining allows for the noninvasive tracing of fungal colonisation along the root system over time. We present MycoRed, a useful innovative method that will expand and complement currently used fungal visualisation techniques to advance knowledge in the field of AM symbiosis.

Alkaline extract of the seaweed Ascophyllum nodosum stimulates arbuscular mycorrhizal fungi and their endomycorrhization of plant roots.

Ascophyllum nodosum extracts (ANE) are well-established plant biostimulants that improve stress tolerance and crop vigour, while also having been shown to stimulate soil microbes. The intersection of these two stimulatory activities, and how they combine to enhance plant health, however, remains poorly understood. In the present study, we aimed to evaluate: (1) the direct effect of ANE on the arbuscular mycorrhizal fungus Rhizophagus irregularis, and (2) whether ANE influences endomycorrhization in plants. ANE enhanced development of R. irregularis in vitro, showing greater spore germination, germ tube length, and hyphal branching. Greenhouse-grown Medicago truncatula drench-treated with ANE formed mycorrhizal associations faster (3.1-fold higher mycorrhization at week 4) and grew larger (29% greater leaf area by week 8) than control plants. Foliar applications of ANE also increased root colonization and arbuscular maturity, but did not appear to enhance plant growth. Nonetheless, following either foliar or drench application, M. truncatula genes associated with establishment of mycorrhizae were expressed at significantly higher levels compared to controls. These results suggest that ANE enhances mycorrhization through both direct stimulation of arbuscular mycorrhizal fungus growth and through stimulation of the plant's accommodation of the symbiont, together promoting the establishment of this agriculturally vital plant-microbe symbiosis.

Target of rapamycin, PvTOR, is a key regulator of arbuscule development during mycorrhizal symbiosis in Phaseolus.

Target of rapamycin (TOR) is a conserved central growth regulator in eukaryotes that has a key role in maintaining cellular nutrient and energy status. Arbuscular mycorrhizal (AM) fungi are mutualistic symbionts that assist the plant in increasing nutrient absorption from the rhizosphere. However, the role of legume TOR in AM fungal symbiosis development has not been investigated. In this study, we examined the function of legume TOR in the development and formation of AM fungal symbiosis. RNA-interference-mediated knockdown of TOR transcripts in common bean (Phaseolus vulgaris) hairy roots notably suppressed AM fungus-induced lateral root formation by altering the expression of root meristem regulatory genes, i.e., UPB1, RGFs, and sulfur assimilation and S-phase genes. Mycorrhized PvTOR-knockdown roots had significantly more extraradical hyphae and hyphopodia than the control (empty vector) roots. Strong promoter activity of PvTOR was observed at the site of hyphal penetration and colonization. Colonization along the root length was affected in mycorrhized PvTOR-knockdown roots and the arbuscules were stunted. Furthermore, the expression of genes induced by AM symbiosis such as SWEET1, VPY, VAMP713, and STR was repressed under mycorrhized conditions in PvTOR-knockdown roots. Based on these observations, we conclude that PvTOR is a key player in regulating arbuscule development during AM symbiosis in P. vulgaris. These results provide insight into legume TOR as a potential regulatory factor influencing the symbiotic associations of P. vulgaris and other legumes.

Evaluation of the Effect of Strigolactones and Synthetic Analogs on Fungi.

Strigolactones (SLs) are components of root exudates as a consequence of active release from the roots into the soil. Notably, they have been described as stimulants of seed germination in parasitic plants and of the presymbiotic growth in arbuscular mycorrhizal (AM) fungi, which are a crucial component of the plant root beneficial microbiota. SLs have therefore the potential to influence other microbes that proliferate in the soil around the roots and may interact with plants. A direct effect of SL analogs on the in vitro growth of a number of saprotrophic or plant pathogenic fungi was indeed reported.Here we describe a standardized method to evaluate the effect of SLs or their synthetic analogs on AM and filamentous fungi. For AM fungi, we propose a spore germination assay since it is more straightforward than the hyphal branching assay and it does not require deep expertise and skills. For filamentous fungi that can grow in axenic cultures, we describe the assay based on SLs embedded in the solid medium or dissolved in liquid cultures where the fungus is inoculated to evaluate the effect on growth, hyphal branching or conidia germination. These assays are of help to test the activity of natural SLs as well as of newly designed SL analogs for basic and applied research.

Controlled Assays for Phenotyping the Effects of Strigolactone-Like Molecules on Arbuscular Mycorrhiza Development.

Arbuscular mycorrhiza is an ancient symbiosis between most land plants and fungi of the Glomeromycotina, in which the fungi provide mineral nutrients to the plant in exchange for photosynthetically fixed organic carbon. Strigolactones are important signals promoting this symbiosis, as they are exuded by plant roots into the rhizosphere to stimulate activity of the fungi. In addition, the plant karrikin signaling pathway is required for root colonization. Understanding the molecular mechanisms underpinning root colonization by AM fungi, requires the use of plant mutants as well as treatments with different environmental conditions or signaling compounds in standardized cocultivation systems to allow for reproducible root colonization phenotypes. Here we describe how we set up and quantify arbuscular mycorrhiza in the model plants Lotus japonicus and Brachypodium distachyon under controlled conditions. We illustrate a setup for open pot culture as well as for closed plant tissue culture (PTC) containers, for plant-fungal cocultivation in sterile conditions. Furthermore, we explain how to harvest, store, stain, and image AM roots for phenotyping and quantification of different AM structures.

A coumarin exudation pathway mitigates arbuscular mycorrhizal incompatibility in Arabidopsis thaliana.

KEY MESSAGE: Overexpression of genes involved in coumarin production and secretion can mitigate mycorrhizal incompatibility in nonhost Arabidopsis plants. The coumarin scopoletin, in particular, stimulates pre-penetration development and metabolism in mycorrhizal fungi. Although most plants can benefit from mutualistic associations with arbuscular mycorrhizal (AM) fungi, nonhost plant species such as the model Arabidopsis thaliana have acquired incompatibility. The transcriptional response of Arabidopsis to colonization by host-supported AM fungi switches from initial AM recognition to defense activation and plant growth antagonism. However, detailed functional information on incompatibility in nonhost-AM fungus interactions is largely missing. We studied interactions between host-sustained AM fungal networks of Rhizophagus irregularis and 18 Arabidopsis genotypes affected in nonhost penetration resistance, coumarin production and secretion, and defense (salicylic acid, jasmonic acid, and ethylene) and growth hormones (auxin, brassinosteroid, cytokinin, and gibberellin). We demonstrated that root-secreted coumarins can mitigate incompatibility by stimulating fungal metabolism and promoting initial steps of AM colonization. Moreover, we provide evidence that major molecular defenses in Arabidopsis do not operate as primary mechanisms of AM incompatibility nor of growth antagonism. Our study reveals that, although incompatible, nonhost plants can harbor hidden tools that promote initial steps of AM colonization. Moreover, it uncovered the coumarin scopoletin as a novel signal in the pre-penetration dialogue, with possible implications for the chemical communication in plant-mycorrhizal fungi associations.

Propagation and characterization of viable arbuscular mycorrhizal fungal spores within maize plant (Zea mays L.).

BACKGROUND: The harmful effect of chemical fertilizer application on human health and the environment as a modern method of meeting the food demand of the increasing world population demands an urgent alternative that is environmentally friendly, which will pose no harm to human health and the environment. Arbuscular mycorrhizal fungi (AMF) are beneficial soil microorganisms that provide various ecological functions in increasing soil fertility and enhancing plant growth. This present study aimed to propagate, characterize and examine the effect of viable arbuscular mycorrhizal fungal spores on maize (Zea mays L) hosts using molecular methods. The propagation of AMF in the host plant using sterile soil and vermiculite was conducted in the greenhouse. RESULT: The effect of AMF inoculation revealed a significant difference (P > 0.05) in maize growth, root colonization and AMF spore count when compared with the control. In all the parameters measured in this study, all the AMF spores propagated had a positive effect on the maize plant over the control, with the highest value mostly recorded in Rhizophagus irregularis AOB1. The molecular characterization of the spore using a specific universal primer for Glomeromycota established the success of the propagation process, which enhanced the classification of the AMF species into Rhizophagus irregularis OAB1, Glomus mosseae OAB2 and Paraglomus occultum OAB3. CONCLUSION: This finding will be a starting point in producing arbuscular mycorrhizal inoculum as a biofertilizer to enhance plant growth promotion. © 2021 Society of Chemical Industry.

Differences in aluminum tolerance and immobilization between two indigenous ectomycorrhizal fungi Lactarius deliciosus and Pisolithus tinctorius from Southwest China's forest stands.

Aluminum (Al) toxicity severely decreases plant growth and productivity in acidic soil globally. Ectomycorrhizal (ECM) fungi can promote host plant's Al-tolerance by acting as a physical barrier or bio-filter. However, little information is available on the role of ECM fungus on Al immobilization with respect to Al-tolerance. This present study aimed to screen a promising indigenous ECM fungus with high Al-tolerance and to understand its role in Al immobilization related to Al-tolerance. Two ECM fungal strains (Lactarius deliciosus 2 and Pisolithus tinctorius 715) isolated from forest stands in Southwest China were cultured in vitro with 0.0, 1.0 or 2.0 mM Al addition for 21 days to compare their Al accumulation and Al-tolerance. Meanwhile, fungal mycelia were incubated in 0.037 mM Al3+ solutions, and then Al3+ concentrations in the solution were determined at time 2, 5, 10, 20, 40, 60, 120, 180, and 240 min, and the Al3+ immobilization characteristics were evaluated using the pseudo-first order, pseudo-second order and intraparticle diffusion models. Results showed that 1.0 or 2.0 mM Al3+ addition significantly increased fungal biomass production by 23% or 41% in L. deliciosus 2, not in P. tinctorius 715. Fungal Al3+ concentrations in L. deliciosus 2 and P. tinctorius 715 were significantly increased by 293% and 103% under 2.0 mM than under 1.0 mM Al3+ addition. The pH values in the culture solution were significantly decreased by 0.43 after 21 d fungus growth but no changes between these two fungi under the same Al3+ addition. Fungal Al3+ immobilization showed a three-stage trend with initially a rapid rate followed a relatively slower rate until reaching equilibrium. The pseudo-second order model was the best (R2 = 0.98 and 0.99 for L. deliciosus 2 and P. tinctorius 715) to fit the experimentally observed data among the three models. Compared to P. tinctorius 715, L. deliciosus 2 also had greater intercept value, cation exchange capacity (CEC), and extracellular Al3+ proportion in fungal mycelia. Additionally, bio-concentration on Al3+, active site numbers for Al3+, boundary layer thickness, CEC, and immobilization on the cell wall in fungal mycelia were involved in ECM fungal Al-tolerance. These results show that both ECM fungi are Al-tolerant while L. deliciosus 2 is a promising indigenous ECM isolate with higher Al-tolerance in Southwest China, and they can be hence applied to the afforestation and ecological restoration in acidic soil.

Insufficient sampling constrains our characterization of plant microbiomes.

Plants host diverse microbial communities, but there is little consensus on how we sample these communities, and this has unknown consequences. Using root and leaf tissue from showy milkweed (Asclepias speciosa), we compared two common sampling strategies: (1) homogenizing after subsampling (30 mg), and (2) homogenizing bulk tissue before subsampling (30 mg). We targeted bacteria, arbuscular mycorrhizal (AM) fungi and non-AM fungi in roots, and foliar fungal endophytes (FFE) in leaves. We further extracted DNA from all of the leaf tissue collected to determine the extent of undersampling of FFE, and sampled FFE twice across the season using strategy one to assess temporal dynamics. All microbial groups except AM fungi differed in composition between the two sampling strategies. Community overlap increased when rare taxa were removed, but FFE and bacterial communities still differed between strategies, with largely non-overlapping communities within individual plants. Increasing the extraction mass 10 × increased FFE richness ~ 10 ×, confirming the severe undersampling indicated in the sampling comparisons. Still, seasonal patterns in FFEs were apparent, suggesting that strong drivers are identified despite severe undersampling. Our findings highlight that current sampling practices poorly characterize many microbial groups, and increased sampling intensity is necessary for increase reproducibility and to identify subtler patterns in microbial distributions.

Extraction of short chain chitooligosaccharides from fungal biomass and their use as promoters of arbuscular mycorrhizal symbiosis.

Short chain chitooligosaccharides (COs) are chitin derivative molecules involved in plant-fungus signaling during arbuscular mycorrhizal (AM) interactions. In host plants, COs activate a symbiotic signalling pathway that regulates AM-related gene expression. Furthermore, exogenous CO application was shown to promote AM establishment, with a major interest for agricultural applications of AM fungi as biofertilizers. Currently, the main source of commercial COs is from the shrimp processing industry, but purification costs and environmental concerns limit the convenience of this approach. In an attempt to find a low cost and low impact alternative, this work aimed to isolate, characterize and test the bioactivity of COs from selected strains of phylogenetically distant filamentous fungi: Pleurotus ostreatus, Cunninghamella bertholletiae and Trichoderma viride. Our optimized protocol successfully isolated short chain COs from lyophilized fungal biomass. Fungal COs were more acetylated and displayed a higher biological activity compared to shrimp-derived COs, a feature that-alongside low production costs-opens promising perspectives for the large scale use of COs in agriculture.

Arbuscular mycorrhizal enhancement of phosphorus uptake and yields of maize under high planting density in the black soil region of China.

Arbuscular mycorrhizal (AM) symbioses are an attractive means of improving the efficiency of soil phosphorus (P) that difficult to be used by plants and may provide a sustainable way of maintaining high yields while reducing P applications. However, quantifying the contribution of indigenous AM fungi on phosphorus uptake and yields of maize (Zea mays L.) under field conditions is not particularly clear. Mesh-barrier compartments were applied to monitor the distribution of hyphal P uptake throughout the experimental period under different planting densities and soil depths, over two consecutive years. AM symbioses enhanced plant P-acquisition efficiency, especially during the silking stage, and hyphae of AM fungi was assessed to contribution 19.4% at most to total available P content of soil. Moreover, the pattern of AM depletion of soil P generally matched shoot nutrient demand under the high planting density, which resulted in significantly increased yield in 2014. Although the hyphal length density was significantly decreased with soil depth, AM fungi still had high potential for P supply in deeper soil. It demonstrates the great potential of indigenous AM fungi to maize productivity in the high-yield area of China, and it would further provide the possibility of elimination P fertilizer applications to maintain high yields.

Selection of suitable reference genes for quantitative real time PCR in different Tulasnella isolates and orchid-fungus symbiotic germination system.

Under natural conditions, mycorrhizal symbiosis accompanies nearly the entire life cycle of orchids from seed germination through to flowering and fruiting. Tulasnella-like orchid mycorrhizal fungi are the most common mycorrhizal fungi found in association with orchid species. Presently suitable reference genes have not been systematically selected for the quantification of gene expression via Real-Time Quantitative Reverse Transcription PCR (RT-qPCR). We evaluated 12 candidate Tulasnella genes in nine different Tulasnella isolates and in the Dendrobium-fungal symbiotic germination associations followed by statistical analysis using the programs Bestkeeper, geNorm, and Normfinder to analyze the expression stability of the individual genes. The results showed that the EF2, UBC, and PP2A genes had the highest rankings with relatively stable expression levels across the different genotypes and during the symbiotic seed germination process by the three programs, and may be suitable for RT-qPCR normalization. Furthermore, the gene encoding C-5 Sterol desaturase (C5SD) was selected to verify the reliability of EF2, UBC, and PP2A expression during the Tulasnella-Dendrobium symbiotic seed germination process. This study is the first systematic exploration of optimal reference genes for gene expression studies during the colonization of orchid seeds by the mycorrhizal fungus Tulasnella.

A Novel Putative Microtubule-Associated Protein Is Involved in Arbuscule Development during Arbuscular Mycorrhiza Formation.

The formation of arbuscular mycorrhizal (AM) symbiosis requires plant root host cells to undergo major structural and functional reprogramming to house the highly branched AM fungal structure for the reciprocal exchange of nutrients. These morphological modifications are associated with cytoskeleton remodelling. However, molecular bases and the role of microtubules (MTs) and actin filament dynamics during AM formation are largely unknown. In this study, the tomato tsb (tomato similar to SB401) gene, belonging to a Solanaceae group of genes encoding MT-associated proteins (MAPs) for pollen development, was found to be highly expressed in root cells containing arbuscules. At earlier stages of mycorrhizal development, tsb overexpression enhanced the formation of highly developed and transcriptionally active arbuscules, while tsb silencing hampers the formation of mature arbuscules and represses arbuscule functionality. However, at later stages of mycorrhizal colonization, tsb overexpressing (OE) roots accumulate fully developed transcriptionally inactive arbuscules, suggesting that the collapse and turnover of arbuscules might be impaired by TSB accumulation. Imaging analysis of the MT cytoskeleton in cortex root cells OE tsb revealed that TSB is involved in MT bundling. Taken together, our results provide unprecedented insights into the role of novel MAP in MT rearrangements throughout the different stages of the arbuscule life cycle.

Higher temperatures and lower annual rainfall do not restrict, directly or indirectly, the mycorrhizal colonization of barley (Hordeum vulgare L.) under rainfed conditions.

Whereas the role of arbuscular mycorrhizal fungi (AMF) in plant growth improvement has been well described in agroecosystems, little is known about the effect of environmental factors on AMF root colonization status of barley, the fourth most important cereal crop all over the world. In order to understand the influence of environmental factors, such as climatic and soil physico-chemical properties, on the spontaneous mycorrhizal ability of barley (Hordeum vulgare L.), a field investigation was conducted in 31 different sites in sub-humid, upper and middle semi-arid areas of Northern Tunisia. Mycorrhizal root colonization of H. vulgare varied considerably among sites. Principal component analysis showed that barley mycorrhization is influenced by both climatic and edaphic factors. A partial least square structural equation modelling (PLS-SEM) revealed that 39% (R²) of the total variation in AMF mycorrhizal rate of barley roots was mainly explained by chemical soil properties and climatic characteristics. Whereas barley root mycorrhizal rates were inversely correlated with soil organic nitrogen (ON), available phosphorus amounts (P), altitude (Z), average annual rainfall (AAR), they were directly correlated with soil pH and temperature. Our results indicated that AMF root colonization of barley was strongly related to climatic characteristics than chemical soil properties. The current study highlights the importance of the PLS-SEM to understand the interactions between climate, soil properties and AMF symbiosis of barley in field conditions.