Asymbiotic mass production of the arbuscular mycorrhizal fungus Rhizophagus clarus

Arbuscular mycorrhizal (AM) symbiosis is a mutually beneficial interaction between fungi and land plants and promotes global phosphate cycling in terrestrial ecosystems. AM fungi are recognised as obligate symbionts that require root colonisation to complete a life cycle involving the production of propagules, asexual spores. Recently, it has been shown that Rhizophagus irregularis can produce infection-competent secondary spores asymbiotically by adding a fatty acid, palmitoleic acid. Furthermore, asymbiotic growth can be supported using myristate as a carbon and energy source for their asymbiotic growth to increase fungal biomass. However, the spore production and the ability of these spores to colonise host roots were still limited compared to the co-culture of the fungus with plant roots. Here we show that a combination of two plant hormones, strigolactone and jasmonate, induces the production of a large number of infection-competent spores in asymbiotic cultures of Rhizophagus clarus HR1 in the presence of myristate and organic nitrogen. Inoculation of asymbiotically-generated spores promoted the growth of host plants, as observed for spores produced by symbiotic culture system. Our findings provide a foundation for the elucidation of hormonal control of the fungal life cycle and the development of inoculum production schemes.

Conservation and Diversity in Gibberellin-Mediated Transcriptional Responses Among Host Plants Forming Distinct Arbuscular Mycorrhizal Morphotypes

Morphotypes of arbuscular mycorrhizal (AM) symbiosis, Arum, Paris, and Intermediate types, are mainly determined by host plant lineages. It was reported that the phytohormone gibberellin (GA) inhibits the establishment of Arum-type AM symbiosis in legume plants. In contrast, we previously reported that GA promotes the establishment of Paris-type AM symbiosis in Eustoma grandiflorum, while suppressing Arum-type AM symbiosis in a legume model plant, Lotus japonicus. This raises a hitherto unexplored possibility that GA-mediated transcriptional reprogramming during AM symbiosis is different among plant lineages as the AM morphotypes are distinct. Here, our comparative transcriptomics revealed that several symbiosis-related genes were commonly upregulated upon AM fungal colonization in L. japonicus (Arum-type), Daucus carota (Intermediate-type), and E. grandiflorum (Paris-type). Despite of the similarities, the fungal colonization levels and the expression of symbiosis-related genes were suppressed in L. japonicus and D. carota but were promoted in E. grandiflorum in the presence of GA. Moreover, exogenous GA inhibited the expression of genes involved in biosynthetic process of the pre-symbiotic signal component, strigolactone, which resulted in the reduction of its endogenous accumulation in L. japonicus and E. grandiflorum. Additionally, differential regulation of genes involved in sugar metabolism suggested that disaccharides metabolized in AM roots would be different between L. japonicus and D. carota/E. grandiflorum. Therefore, this study uncovered the conserved transcriptional responses during mycorrhization regardless of the distinct AM morphotype. Meanwhile, we also found diverse responses to GA among phylogenetically distant AM host plants.

Brassinosteroids Benefit Plants Performance by Augmenting Arbuscular Mycorrhizal Symbiosis

Brassinosteroids (BR) and Arbuscular mycorrhizas (AM) symbiosis play an important role in improving plant growth and development. Previous studies have shown that there is a complex regulatory network between phytohormones and AM symbiosis.

Sugar transporters of the SWEET family and their role in arbuscular mycorrhiza

Plant sugar transporters play an essential role in the organism’s productivity by carrying out carbohydrate transportation from source cells in the leaves to sink cells in the cortex. In addition, they aid in the regulation of a substantial part of the exchange of nutrients with microorganisms in the rhizosphere (bacteria and fungi), an activity essential to the formation of symbiotic relationships. This review pays special attention to carbohydrate nutrition during the development of arbuscular mycorrhiza (AM), a symbiosis of plants with fungi from the Glomeromycotina subdivision. This relationship results in the host plant receiving micronutrients from the mycosymbiont, mainly phosphorus, and the fungus receiving carbon assimilation products in return. While the efficient nutrient transport pathways in AM symbiosis are yet to be discovered, SWEET sugar transporters are one of the three key families of plant carbohydrate transporters. Specific AM symbiosis transporters can be identified among the SWEET proteins. The survey provides data on the study history, structure and localization, phylogeny and functions of the SWEET proteins. A high variability of both the SWEET proteins themselves and their functions is noted along with the fact that the same proteins may perform different functions in different plants. A special role is given to the SWEET transporters in AM development. SWEET transporters can also play a key role in abiotic stress tolerance, thus allowing plants to adapt to adverse environmental conditions. The development of knowledge about symbiotic systems will contribute to the creation of microbial preparations for use in agriculture in the Russian Federation.

Mycorrhiza-Induced Alterations in Metabolome of Medicago lupulina Leaves during Symbiosis Development

The present study is aimed at disclosing metabolic profile alterations in the leaves of the Medicago lupulina MlS-1 line that result from high-efficiency arbuscular mycorrhiza (АМ) symbiosis formed with Rhizophagus irregularis under condition of a low phosphorus level in the substrate. A highly effective AM symbiosis was established in the period from the stooling to the shoot branching initiation stage (the efficiency in stem height exceeded 200%). Mycorrhization led to a more intensive accumulation of phosphates (glycerophosphoglycerol and inorganic phosphate) in M. lupulina leaves. Metabolic spectra were detected with GS-MS analysis. The application of complex mathematical analyses made it possible to identify the clustering of various groups of 320 metabolites and thus demonstrate the central importance of the carbohydrate and carboxylate-amino acid clusters. The results obtained indicate a delay in the metabolic development of mycorrhized plants. Thus, AM not only accelerates the transition between plant developmental stages but delays biochemical “maturation” mainly in the form of a lag of sugar accumulation in comparison with non-mycorrhized plants. Several methods of statistical modeling proved that, at least with respect to determining the metabolic status of host-plant leaves, stages of phenological development have priority over calendar age.

Medicago SPX1 and SPX3 regulate phosphate homeostasis, mycorrhizal colonization, and arbuscule degradation

To acquire sufficient mineral nutrients such as phosphate (Pi) from the soil, most plants engage in symbiosis with arbuscular mycorrhizal (AM) fungi. Attracted by plant-secreted strigolactones (SLs), the fungi colonize the roots and form highly branched hyphal structures called arbuscules inside inner cortex cells. The host plant must control the different steps of this interaction to maintain its symbiotic nature. However, how plants sense the amount of Pi obtained from the fungus, and how this determines the arbuscule lifespan, are far from understood. Here, we show that Medicago truncatula SPX-domain containing proteins SPX1 and SPX3 regulate root Pi starvation responses, in part by interacting with PHOSPHATE RESPONSE REGULATOR2, as well as fungal colonization and arbuscule degradation. SPX1 and SPX3 are induced upon Pi starvation but become more restricted to arbuscule-containing cells upon the establishment of symbiosis. This induction in arbuscule-containing cells is associated with the presence of cis-regulatory AW-boxes and transcriptional regulation by the WRINKLED1-like transcription factor WRI5a. Under Pi-limiting conditions, SPX1 and SPX3 facilitate the expression of the SL biosynthesis gene DWARF27, which could help explain the increased fungal branching in response to root exudates. Later, in arbuscule-containing cells, SPX1 and SPX3 redundantly control arbuscule degradation. Thus, SPX proteins play important roles as phosphate sensors to maintain a beneficial AM symbiosis.

An Ancestral Function of Strigolactones as Symbiotic Rhizosphere Signals

In flowering plants, carotenoid-derived strigolactones (SLs) have dual functions as hormones that regulate growth and development, and as rhizosphere signaling molecules that induce symbiosis with arbuscular mycorrhizal (AM) fungi. Here, we report the identification of bryosymbiol (BSB), a previously unidentified SL from the bryophyte Marchantia paleacea. BSB is also found in vascular plants, indicating that it is ancestral in land plants. BSB synthesis is enhanced at AM symbiosis permissive conditions and BSB deficient mutants are impaired in AM symbiosis. In contrast, the absence of BSB synthesis has little effect on the growth and gene expression. We show that the introduction of the SL receptor of Arabidopsis renders M. paleacea cells BSB-responsive. These results suggest that BSB is not perceived by M. paleacea cells due to the lack of cognate SL receptors. We propose that SLs originated as AM symbiosis-inducing rhizosphere signaling molecules and were later recruited as plant hormone.

The Synergy of Arbuscular Mycorrhizal Fungi and Exogenous Abscisic Acid Benefits Robinia pseudoacacia L. Growth through Altering the Distribution of Zn and Endogenous Abscisic Acid

The simultaneous effects of arbuscular mycorrhizal (AM) fungi and abscisic acid (ABA) on the tolerance of plants to heavy metal (HM) remain unclear. A pot experiment was carried out to clarify the effects of simultaneous applications of AM fungi and ABA on plant growth, Zn accumulation, endogenous ABA contents, proline metabolism, and the oxidative injury of black locust (Robinia pseudoacacia L.) exposed to excess Zn stress. The results suggested that exogenously applied ABA positively enhanced AM colonization, and that the growth of plants only with AM fungi was improved by ABA application. Under Zn stress, AM inoculation and ABA application increased the ABA content in the root/leaf (increased by 48–172% and 92%, respectively) and Zn content in the root/shoot (increased by 63–152% and 61%, respectively) in AM plants, but no similar trends were observed in NM plants. Additionally, exogenous ABA addition increased the proline contents of NM roots concomitantly with the activities of the related synthases, whereas it reduced the proline contents and the activity of Δ1-pyrroline-5-carboxylate synthetase in AM roots. Under Zn stress, AM inoculation and ABA application decreased H2O2 contents and the production rate of O2, to varying degrees. Furthermore, in the roots exposed to Zn stress, AM inoculation augmented the activities of SOD, CAT, POD and APX, and exogenously applied ABA increased the activities of SOD and POD. Overall, AM inoculation combined with ABA application might be beneficial to the survival of black locust under Zn stress by improving AM symbiosis, inhibiting the transport of Zn from the roots to the shoots, increasing the distribution of ABA in roots, and stimulating antioxidant defense systems.

Production of Organic Acids by Arbuscular Mycorrhizal Fungi and Their Contribution in the Mobilization of Phosphorus Bound to Iron Oxides

Most plants living in tropical acid soils depend on the arbuscular mycorrhizal (AM) symbiosis for mobilizing low-accessible phosphorus (P), due to its strong bonding by iron (Fe) oxides. The roots release low-molecular-weight organic acids (LMWOAs) as a mechanism to increase soil P availability by ligand exchange or dissolution. However, little is known on the LMWOA production by AM fungi (AMF), since most studies conducted on AM plants do not discriminate on the LMWOA origin. This study aimed to determine whether AMF release significant amounts of LMWOAs to liberate P bound to Fe oxides, which is otherwise unavailable for the plant. Solanum lycopersicum L. plants mycorrhized with Rhizophagus irregularis were placed in a bicompartmental mesocosm, with P sources only accessible by AMF. Fingerprinting of LMWOAs in compartments containing free and goethite-bound orthophosphate (OP or GOE-OP) and phytic acid (PA or GOE-PA) was done. To assess P mobilization via AM symbiosis, P content, photosynthesis, and the degree of mycorrhization were determined in the plant; whereas, AM hyphae abundance was determined using lipid biomarkers. The results showing a higher shoot P content, along with a lower N:P ratio and a higher photosynthetic capacity, may be indicative of a higher photosynthetic P-use efficiency, when AM plants mobilized P from less-accessible sources. The presence of mono-, di-, and tricarboxylic LMWOAs in compartments containing OP or GOE-OP and phytic acid (PA or GOE-PA) points toward the occurrence of reductive dissolution and ligand exchange/dissolution reactions. Furthermore, hyphae grown in goethite loaded with OP and PA exhibited an increased content of unsaturated lipids, pointing to an increased membrane fluidity in order to maintain optimal hyphal functionality and facilitate the incorporation of P. Our results underpin the centrality of AM symbiosis in soil biogeochemical processes, by highlighting the ability of the AMF and accompanying microbiota in releasing significant amounts of LMWOAs to mobilize P bound to Fe oxides.

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.