G3 PhyloChip Analysis Confirms the Promise of Plant-Based Culture Media for Unlocking the Composition and Diversity of the Maize Root Microbiome and for Recovering Unculturable Candidate Divisions/Phyla

Abstract

The rapid development of high-throughput techniques and expansion of bacterial databases have accelerated efforts to bring plant microbiomes into cultivation. We introduced plant-only-based culture media as a successful candidate to mimic the nutritional matrices of plant roots. We herein employed a G3 PhyloChip microarray to meticulously characterize the culture-dependent and -independent bacterial communities of the maize root compartments, the endo- and ecto-rhizospheres. An emphasis was placed on the preference of the growth of unculturable candidate divisions/phyla on plant-only-based culture media over standard culture media (nutrient agar). A total of 1,818 different operational taxonomic units (OTUs) were resolved representing 67 bacterial phyla. Plant-only-based culture media displayed particular affinity towards recovering endophytic over ectophytic rhizobacteria. This was shown by the slightly higher recovery of CFUs for endophytes on plant-only-based culture media (26%) than on standard culture media (10%) as well as the higher taxa richness and numbers of exclusive families of unculturable divisions/phyla. Out of 30 bacterial phyla (comprising >95% of the whole population), 13 were of a significantly higher incidence on plant-only-based culture media, 6 phyla of which were not-yet-cultured (Atribacteria, OP9; Dependentiae, TM6; Latescibacteria, WS3; Marinimicrobia, SAR406; Omnitrophica, OP3; BRC1). Furthermore, plant-only-based culture media significantly enriched less abundant and/or hard-to-culture bacterial phyla (Acidobacteria, Gemmatimonadetes, and Tenericutes). These results present conclusive evidence of the ability of plant-only-based culture media to bring the plant-fed in situ microbiome into the status of plant-fed in vitro cultures, and to widen the scope of cultivation of heretofore-unculturable bacterial divisions/phyla.

Culturomics of the plant prokaryotic microbiome and the dawn of plant-based culture media – A review

“…They contain copious sources of nutritional macromolecules, proteins and carbohydrates, major and minor elements, amino acids and vitamins: a composition that is nearly impossible to tailor in one single or a general synthetic culture medium recommended for common cultivation of the plant microbiota that are used to enjoy such in situ nutritional milieu. Therefore, serious efforts were made to introduce and research natural culture media based on the plant, and its inhabiting microbiota, as a sole source of nutrients, in the form of juices, saps and/or dehydrated powders [80], [81], [82], [83], [84], [85], [86], [87] (Fig. 2).…”

“…[82]Lucerne shoots powderSoleIsolatedLucerne roots Pedobacter sp. [82]Lucerne shoots powderSoleIsolatedLucerne rootsVerrucomicrobia Subdivision 1 [88]Potato root extractsSupplementIsolatedPotato roots Paenibacillus gorilla [6]Mango juiceSoleIsolatedGorilla stool Paenibacillus camerounensis [6]Mango juiceSoleIsolatedGorilla stool Oenococcus oeni [89]Tomato juiceSupplementIsolatedFermented wines Rhizobacter daucus [90]Potato extractSupplementIsolatedCarrot rootsBRC1 [85]Clover shoot powderSoleEnrichedMaize rootsGracilibacteria (GN02) [85]Clover shoot powderSoleEnrichedMaize rootsOmnitrophica (OP3) [85]Clover shoot powderSoleEnrichedMaize rootsAtribacteria (OP9) [85]Clover shoot powderSoleEnrichedMaize rootsMarinimicrobia (SAR406) [85]Clover shoot powderSoleEnrichedMaize rootsDependentiae (TM6) [85]Clover shoot powderSoleEnrichedMaize rootsLatescibacteria (WS3) [85]Clover shoot powderSoleEnrichedMaize rootsArmatimonadetes (OP10) [91]Reed plant roots extractSupplementIsolatedReed plant rootsaNumbers between brackets refer to references related.

Fig. 3Plant-based culture media.

…”

An inoculum-dependent culturing strategy (IDC) for the cultivation of environmental microbiomes and the isolation of novel endophytic Actinobacteria

“…To gain insights into their unknown functions and exploit their potentials, different approaches were recently introduced to culture such not-yet-cultured bacteria [2,3]. In this respect, the plantonly-based culture media were presented as natural culture media to replace myriad formulas of synthetic culture media, and strongly recommended to increase the cultivability of the plant microbiota [4][5][6][7][8][9][10]. To alleviate the stress of disproportionate nutrients, present in common culture media, we aimed at culturing maize and sunflower microbiota on the natural nutrients present in the plant inoculum itself, compared to highly diluted plant-only-based culture media and standard R2A.…”

“In situ similis” Culturing of Plant Microbiota: A Novel Simulated Environmental Method Based on Plant Leaf Blades as Nutritional Pads

“…The strategy recovered 13 genera representing the three major phyla of Firmicutes, Proteobacteria, and Actinobacteria. Representatives of these phyla are commonly reported in different compartments of plants subjected to various environments, including both biotic and abiotic stresses (Koberl et al, 2011;Marasco et al, 2012;Sarhan et al, 2016Sarhan et al, , 2018Hegazi et al, 2017).…”