Bacteria Associated With a Commercial Mycorrhizal Inoculum: Community Composition and Multifunctional Activity as Assessed by Illumina Sequencing and Culture-Dependent Tools

Abstract

The implementation of sustainable agriculture encompasses practices enhancing the activity of beneficial soil microorganisms, able to modulate biogeochemical soil cycles and to affect soil fertility. Among them, arbuscular mycorrhizal fungi (AMF) establish symbioses with the roots of most food crops and play a key role in nutrient uptake and plant protection from biotic and abiotic stresses. Such beneficial services, encompassing improved crop performances, and soil resources availability, are the outcome of the synergistic action of AMF and the vast communities of mycorrhizospheric bacteria living strictly associated with their mycelium and spores, most of which showing plant growth promoting (PGP) activities, such as the ability to solubilize phosphate and produce siderophores and indole acetic acid (IAA). One of the strategies devised to exploit AMF benefits is represented by the inoculation of selected isolates, either as single species or in a mixture. Here, for the first time, the microbiota associated with a commercial AMF inoculum was identified and characterized, using a polyphasic approach, i.e., a combination of culture-dependent analyses and metagenomic sequencing. Overall, 276 bacterial genera were identified by Illumina high-throughput sequencing, belonging to 165 families, 107 orders, and 23 phyla, mostly represented by Proteobacteria and Bacteroidetes. The commercial inoculum harbored a rich culturable heterotrophic bacterial community, whose populations ranged from 2.5 to 6.1 × 10⁶ CFU/mL. The isolation of functional groups allowed the selection of 36 bacterial strains showing PGP activities. Among them, 14 strains showed strong IAA and/or siderophores production and were affiliated with Actinomycetales (Microbacterium trichotecenolyticum, Streptomyces deccanensis/scabiei), Bacillales (Bacillus litoralis, Bacillus megaterium), Enterobacteriales (Enterobacter), Rhizobiales (Rhizobium radiobacter). This work demonstrates for the first time that an AMF inoculum, obtained following industrial production processes, is home of a large and diverse community of bacteria with important functional PGP traits, possibly acting in synergy with AMF and providing additional services and benefits. Such bacteria, available in pure culture, could be utilized, individually and/or in multispecies consortia with AMF, as biofertilizers and bioenhancers in sustainable agroecosystems, aimed at minimizing the use of chemical fertilizers and pesticides, promoting primary production, and maintaining soil health and fertility.

Keywords: arbuscular mycorrhizal symbionts; indole acetic acid production; metagenomics; mycorrhizosphere; plant-growth promoting bacteria; siderophores production.

Figure 1

Relative abundance of bacterial phyla and classes associated with a commercial AMF inoculum.

Figure 2

Relative abundance of bacterial genera associated with a commercial AMF inoculum.

Figure 3

Affiliation of the sequences of the 14 bacterial strains showing the best PGP traits with the existing 16S rRNA gene sequences. Phylogenetic analysis was inferred by using the Neighbor-Joining method. The evolutionary distances were computed using the Tamura 3-parameter method. The rate variation among sites was modeled with a gamma distribution (shape parameter = 1). Bootstrap (1,000 replicates) values below 70 are not shown. Evolutionary analyses were conducted in MEGA10.