Allocation of Carbon from an Arbuscular Mycorrhizal Fungus, Gigaspora margarita, to Its Gram-Negative and Positive Endobacteria Revealed by High-Resolution Secondary Ion Mass Spectrometry

Arbuscular mycorrhizal fungi are obligate symbionts of land plants; furthermore, some of the species harbor endobacteria. Although the molecular approach increased our knowledge of the diversity and origin of the endosymbiosis and its metabolic possibilities, experiments to address the functions of the fungal host have been limited. In this study, a C flow of the fungus to the bacteria was investigated. Onion seedlings colonized with Gigaspora margarita, possessing Candidatus Glomeribacter gigasporarum (CaGg, Gram-negative, resides in vacuole) and Candidatus Moeniiplasma glomeromycotorum (CaMg, Gram-positive, resides in the cytoplasm,) were labelled with 13CO2. The 13C localization within the mycorrhiza was analyzed using high-resolution secondary ion mass spectrometry (SIMS). Correlative TEM-SIMS analysis of the fungal cells revealed that the 13C/12C ratio of CaGg was the lowest among CaMg and mitochondria and was the highest in the cytoplasm. By contrast, the plant cells, mitochondria, plastids, and fungal cytoplasm, which are contributors to the host, showed significantly higher 13C enrichment than the host cytoplasm. The C allocation patterns implied that CaMg has a greater impact than CaGg on G. margarita, but both seemed to be less burdensome to the host fungus in terms of C cost.

Estabelecimento de Anadenanthera colubrina var. cebil sob inoculação micorrízica e fertilização no Pantanal brasileiro

Este trabalho objetivou avaliar o estabelecimento de Anadenanthera colubrina (Vell.) Brenan var. cebil [Griseb.] Altschul sob inoculação micorrízica e fertilização no Pantanal da Nhecolândia, MS. O experimento foi implantado em março de 2015 em ambiente de cordilheira parcialmente desmatada, em blocos ao acaso distribuídos no micro relevo, e parcelas subdivididas. Os principais tratamentos foram constituídos por mudas submetidas ou não (controle) à inoculação mista com Rhizophagus clarus e Gigaspora margarita, e aos tratamentos secundários de adubação com doses de sulfato de amônio e cloreto de potássio. Entre 2015 e 2017 foram avaliados a abertura do dossel das árvores remanescentes, o percentual de sobrevivência e o crescimento de A. colubrina. Melhores condições de sobrevivência foram verificadas na porção da cordilheira mais próxima à baía, sob sombreamento moderado e com a aplicação, no plantio, de 15 g de sulfato de amônio e de 5 g de cloreto de potássio por muda. A inoculação micorrízica promoveu incremento de 16% em diâmetro do colo aos 23 meses de idade, não aumentando a sobrevivência da espécie. Estudos complementares são necessários para subsidiar estratégias de manejo de A. colubrina, em condições de plantio no Pantanal da Nhecolândia.

PRODUCTION OF SEEDLINGS OF COLUBRINA GLANDULOSA PERKINS WITH DRILLING WASTE FROM OIL WELLS AND MYCORRHIZAL INOCULATION

Oil exploration, whether onshore or offshore, results in residues from the drilling of wells, called gravel. The use of this environmental liability in the production of seedlings for forest restoration could contribute to an appropriate destination for this waste. In the present study, the objective was to evaluate the effect of the substrate formulated with gravel, with and without inoculation with arbuscular mycorrhizal fungi (AMF), on the growth and nutritional status of seedlings of Colubrina glandulosa. The design used was completely randomized in a 5x2 factorial scheme, with five gravel doses (0, 5, 10, 15, and 20%), with and without inoculation with a mixture of AMF spores (Gigaspora margarita¸ Rhizophagus clarus, and Dentiscutata heterogama) with six repetitions, totaling 60 experimental units. Height (H) and stem diameter (SD) were evaluated at 30, 60, 90, and 120 days after seedling transplantation, when the plants were sectioned in shoots (branches + leaves) and roots, to determine dry biomass of shoots (DMS) and of roots (DMR), leaf area index (LAI), rate of length of fine roots colonized by AMF (COL), and chemical composition. The substrate obtained with the application of the lowest gravel dose (5%), without the mycorrhizal inoculation, provided significant increments in H, SD, LAI, DMS, DMR, and COL of the seedlings of Colubrina glandulosa, when compared to the other gravel doses and the presence of mycorrhizal inoculation, 120 days after seedling transplantation.

Tolerance of arbuscular mycorrhizal fungi and microorganisms associated to their hyphosphere to aluminum in soil

Background: The elements that are toxic to arbuscular mycorrhizal fungi (AMF) and soil microorganisms, and the levels at which they affect them, are poorly known. Objective: To quantify the effects of: 1) aluminum added to the soil (0, 50 and 100 mg kg-1) and 2) inoculation with AMF (two isolates without prior exposure to aluminum, Acaulospora delicata and Gigaspora margarita, the native AMF community of a soil contaminated with Al, and a control without AMF) on the development of AMF mycelium and the hyphosphere-associated microbiota. Methods: A system with two compartments was used, a central compartment in which maize was sown and AMF were inoculated and a side compartment without root passage, where only the external mycelium was exposed to the different concentrations of Al. AMF external mycelium exposed to Al was quantified with the fatty-acid biomar-ker 16:1w5. Results and conclusions: Al concentrations up to 100 mg kg-1, controlling the pH of the soil, did not alter mycelium growth of any of the inoculated HMA either in the soil or in the roots. Aluminum added up to 100 mg kg-1 did not have a direct toxic effect on the growth of AMF mycelium and the hyphosphere-associated microbiota.

The Mosaic Architecture of NRPS-PKS in the Arbuscular Mycorrhizal Fungus Gigaspora margarita Shows a Domain With Bacterial Signature

As obligate biotrophic symbionts, arbuscular mycorrhizal fungi (AMF) live in association with most land plants. Among them, Gigaspora margarita has been deeply investigated because of its peculiar features, i.e., the presence of an intracellular microbiota with endobacteria and viruses. The genome sequencing of this fungus revealed the presence of some hybrid non-ribosomal peptide synthases-polyketide synthases (NRPS-PKS) that have been rarely identified in AMF. The aim of this study is to describe the architecture of these NRPS-PKS sequences and to understand whether they are present in other fungal taxa related to G. margarita. A phylogenetic analysis shows that the ketoacyl synthase (KS) domain of one G. margarita NRPS-PKS clusters with prokaryotic sequences. Since horizontal gene transfer (HGT) has often been advocated as a relevant evolutionary mechanism for the spread of secondary metabolite genes, we hypothesized that a similar event could have interested the KS domain of the PKS module. The bacterial endosymbiont of G. margarita, Candidatus Glomeribacter gigasporarum (CaGg), was the first candidate as a donor, since it possesses a large biosynthetic cluster involving an NRPS-PKS. However, bioinformatics analyses do not confirm the hypothesis of a direct HGT from the endobacterium to the fungal host: indeed, endobacterial and fungal sequences show a different evolution and potentially different donors. Lastly, by amplifying a NRPS-PKS conserved fragment and mining the sequenced AMF genomes, we demonstrate that, irrespective of the presence of CaGg, G. margarita, and some other related Gigasporaceae possess such a sequence.

Gigaspora Margarita use to improve flower life in Notocactus and Gymnocalycium plants and roots protection against Fusarium sp.

The paper presents the results of research on cacti such as Gymnocalycium baldianum, Gymnocalycium mihanovichii, Notocactus eugeniae and Notocactus leninghausii, aimed at improving plant growth and defense against the pathogenic fungus Fusarium sp., through the use of Gigaspora margarita inoculated in the growing medium. Objectives of the work were: i) use Gigaspora margarita to assess if the use of arbuscular mycorrhizae can lead to an increase in the growth rate of Notocactus and Gymnocalycium, plants generally slow in their growth cycle; ii) to evaluate if the use of mycorrhizae can lead to an extension of the duration of the flower, to promote pollination of bees; iii) to assess if the use of Gigaspora Margarita allows greater protection of the roots against the fungus Fusarium sp. which often affects the roots of these cacti. The two experimental groups in cultivation were: i) group without arbuscular michorriza, irrigated with water and substrate previously fertilized; group with Gigaspora Margarita, irrigated with water and substrate previously fertilized. All plants treated with Gigaspora margarita showed a significant increase in plant height and circumference, vegetative and root weight of the plants, number of new suckers, number of flowers and fruits and flower life. In addition, there was significant control of the fungus Fusarium sp. in plants in which Gigaspora margarita was inoculated into the substrate, in fact a reduced infection of the roots due to this pathogen was found. The application of mycorrhiza in the cultivation of plants can guarantee the possibility to obtain a higher quality product, a higher resistance to biotic stress, an increase in growth rate, very interesting aspects in the succulent and cactus plants sector.

DAYA INFEKSI DAN EFEKTIVITAS FUNGI MIKORIZA ARBUSKULAR Gigaspora margarita PADA TANAMAN JAGUNG DENGAN MASA SIMPAN YANG BERBEDA

Fungi mikoriza arbuskular (FMA) merupakan salah satu tipe mikoriza yang sangat luas penyebarannya dan membentuk simbiosis mutualisme dengan akar tanaman. Saat ini, pupuk hayati berbahan dasar spora mikoriza sudah banyak dikembangkan, namun laporan tentang berapa lama pupuk ini dapat disimpan tanpa spora kehilangan daya infeksi dan efektivitasnya sangat sedikit. Oleh karena itu, penelitian ini dilakukan dengan tujuan untuk mengevaluasi pengaruh masa simpan terhadap (1) daya infeksi spora Gigaspora margarita dan (2) pertumbuhan tanaman jagung. Spora G. margarita yang telah disimpan selama 0, 6, dan 22 bulan diinokulasikan ke akar kecambah jagung dan ditanam dalam pot bervolume 450 ml berisi media tanam campuran pasir steril dan zeolite (2:1 berdasarkan volume). Infeksi akar diamati pada 1, 2, dan 3 minggu setelah inokulasi FMA. Untuk studi efektivitas, kecambah jagung diinokulasi dengan spora G. margarita dan ditanam dalam polibag yang berisi 8 kg media dan dipelihara di rumah kaca selama 8 minggu. Hasil penelitian menunjukkan bahwa waktu simpan mempengaruhi daya infeksi FMA G. margarita. Spora G. margarita pada semua masa simpan berhasil menginfeksi akar lebih dari 50% 8 minggu setelah inokulasi dan mampu meningkatkan pertumbuhan tanaman jagung.

Gigaspora margarita and Its Endobacterium Modulate Symbiotic Marker Genes in Tomato Roots under Combined Water and Nutrient Stress

As members of the plant microbiota, arbuscular mycorrhizal fungi (AMF) may be effective in enhancing plant resilience to drought, one of the major limiting factors threatening crop productivity. AMF host their own microbiota and previous data demonstrated that endobacteria thriving in Gigaspora margarita modulate fungal antioxidant responses. Here, we used the G. margarita–Candidatus Glomeribacter gigasporarum system to test whether the tripartite interaction between tomato, G. margarita and its endobacteria may improve plant resilience to combined water/nutrient stress. Tomato plants were inoculated with spores containing endobacteria (B+) or not (B-), and exposed to combined water/nutrient stress. Plants traits, AM colonization and expression of AM marker genes were measured. Results showed that mycorrhizal frequency was low and no growth effect was observed. Under control conditions, B+ inoculated plants were more responsive to the symbiosis, as they showed an up-regulation of three AM marker genes involved in phosphate and lipids metabolism compared with B− inoculated or not-inoculated plants. When combined stress was imposed, the difference between fungal strains was still evident for one marker gene. These results indicate that the fungal endobacteria finely modulate plant metabolism, even in the absence of growth response.