Bacterial diversity and composition in rhizosphere and bulk soil
This meta-analysis is based on pairwise data (rhizosphere vs bulk soils) from amplicon sequencing approaches characterizing the taxonomic and functional features. This yielded fundamental insights into crop rhizosphere microbiomes on an across-continent scale and onto the plant-driven microbial taxa and their functional properties in this unique but unified habitat. Our design enabled testing the general influence of crops on their rhizosphere bacterial community across soil and climate environments. This highlights the benefit of using sequencing data in soils to synthesize general microbiome patterns and to indicate specialized functions and life strategies of microbial taxa based on niche differences between rhizosphere and bulk soils.
The fact that rhizosphere microbiota differ from bulk soil microbiota is well documented 11, 15, 26, 27, 28, and this is attributed to significant differences in physico-chemical attributes that result in niche differentiation 4, 20, 29, 30. In addition to the between-niche difference in terms of the environment, the generalized contrast between bulk soil and rhizosphere was the most important source of the distinction in microbiota composition 11, 31. We observed an overall lower bacterial richness, Shannon index, evenness and phylogenetic diversity for the rhizosphere vs bulk soil microbiota in all environments. Consequently, bacterial diversity decreases with increasing substrate availability, the conditions common in the rhizosphere. The general view is that the rhizosphere microbiota, a subset of the community in bulk soil, possess certain similar traits across plants, within varied environments. This underlines the selective effect of the rhizosphere, which to a certain extent has generalized consequences for the rhizobacterial assemblages across crops. This even holds true for bacteria belonging to various classes, orders and families. Though the effect size differs between crop groups (Fig. 1), we stress that, even after accounting for genotypic and environmental differences, the selection of microorganisms common for the rhizosphere still displayed certain similar traits 4, 8, 32.
Of course, the specific environmental conditions fundamentally shift the rhizosphere effect size: the rice rhizosphere, for example, harbors phylogenetically broader bacteria than its corresponding bulk soil. Paddy soils commonly develop anaerobic conditions, and rice plants therefore exhibit a developed aerenchyma. Due to the oxygen release around roots, the Eh and oxygen content in the rice rhizosphere are much higher than in the bulk soil 33, 34 and fluctuate strongly. Thus, a broader phylogeny of both anaerobic and aerobic bacteria colonize the rice rhizosphere. These results indicate that environmental heterogeneity, such as root exudates, Eh and oscillating moisture, interact to shape the rhizosphere selective effect.
The particular microbial taxa recruited to the rhizosphere from the soil microbial reservoir can apparently form a particular core microbiome 11, 13. The core microbiome around roots contributes to plant growth and fitness 16. To date, the core microbiome of plants, regardless of whether in the rhizosphere, endosphere or phyllosphere, has been defined mostly based on taxonomic markers. We, however, emphasize that more attention should be paid to identifying microbes having common functions that are selected for in a general rhizosphere setting. Accordingly, a function-based definition of the microbiome should facilitate efforts to manipulate communities for useful purposes. Our comprehensive analysis revealed a few predominant taxa that are consistently enriched in the rhizosphere, for example the phyla Bacteroidetes and Proteobacteria (Fig. 2). This result underscores the fact that these phyla are generally adapted to C-rich conditions (common in the rhizosphere) 30, 35, and are consequently very similar across diverse plant species. This finding is not surprising because Bacteroidetes and Proteobacteria utilize labile carbon sources for high metabolic activity, fast growth and propagation- They are generally considered to be r-selected, or weedy fast-growing microbiota whose populations fluctuate opportunistically 28. In contrast to the rhizosphere, the bulk soil is generally enriched by other dominant phyla including Acidobacteria (Fig. 2b), which are oligotrophs 36, 37. Interestingly, a few phylum-level taxa are similar between rhizosphere and bulk soil, but finer taxonomic resolution reveals differences (Fig. 2). A case in point is Firmicutes: its class Bacilli is richer in the rhizosphere, whereas the class Clostridia is more abundant in the bulk soils. Similarly, Actinobacteria is richer in the rhizosphere, but its class Thermoleophilia is more abundant in the bulk soils. Consequently, the resulting generalized pattern that emerges based on the selective effect of the rhizosphere depends on the taxonomic resolution and on the fundamental niches at the level of classes and families.
Microbiome formation: from structuring to functions
In microbial networks, highly interconnected species are grouped into a module, within which species interactions are more frequent and intensive than with the remaining community. The rhizosphere bacterial network modularity is higher than that in the bulk soil (Fig. 3 and Table S1). One potential explanation is more pronounced niche differentiations – both spatially and temporally – in the rhizosphere 25 because modules can be interpreted as microbial niches 38, 39. Modularity is one of the main organizing principles of biological networks 40, and the higher modularity in the rhizosphere indicates a more complex topological structure. Nevertheless, the rhizosphere bacterial co-occurrence network is less robust (Fig. 3c). This generalization is not always valid, for example in the case of a single wheat rhizosphere network that was more stable than that in the bulk soil 26. This is attributable to the fact that the networks (Fig. 3) are constructed across plant species. Moreover, that particular rhizosphere is characterized by very high temporal dynamics as compared to the more static conditions in the bulk soil 30, 41. Plant species selectively enrich specific microorganisms by investing in root exudates to feed their rhizosphere microbiota 1, 42. The rhizosphere indigenous microbial community structure often differs remarkably across host species 43. In a soybean cultivation, the microbial community in the rhizosphere was selected via niche filtering, whereas the bulk soil community arose via neutral (stochastic) processes 44. The rhizosphere network allocates more network modules for executive functions, but fewer for network robustness, which partly reflects the fast element cycling in the rhizosphere 45, 46.
Rhizosphere and bulk soil are characterized by different dominant strategies of microbial dormancy: sporulation factors and toxin–antitoxin systems (Fig. 4a). The sporulation factors were more abundant in the bulk soil, whereas the toxin–antitoxin systems were enriched in the rhizosphere (Fig. 4b). During plant growth, roots actively and passively release a broad range of organic compounds into the rhizosphere. These compounds are the driving force for microbial density and activity 29, 47, 48. The sporulation factor was abundant in the bulk dryland soils but played nonsignificant role in paddies (Fig. 4b). Hence, bacterial sporulation is more common in upland soils because the environmental conditions in paddies are more stable and homogeneous, and paddies are always moist. The less role of sporulation played in the rhizosphere (as compared to bulk soil) indirectly confirmed the buffered amplitude of moisture variation, which involves mucilage swelling conditions 49, 50, 51. The genes related to dormancy/sporulation strongly increase with aridity 52. Toxin–antitoxin systems are composed by the genes that encode a toxin protein that inhibits cell growth and an antitoxin that counteracts the toxin 53.
More copiotrophs (e.g. Bacteroidetes and Proteobacteria) inhabited the rhizosphere, as confirmed by their rRNA operon counts, which were considerably higher there (Fig. 4c). Copiotrophs have more operon counts than oligotrophs 54. The lower rRNA operon copy number is common for K-selected microbiota, and leads to slower growth rates and a more stable population. The bacterial functional trait of rRNA operon copy numbers increases with resource availability 22. Organisms with multiple operon counts tend to be r-strategists, which are dominant in resource-abundant conditions and respond more rapidly to nutrient inputs 55, 56, 57. Therefore, copiotrophs are dominant when resources are abundant, such as in the rhizosphere habitat.
Genes related to nitrification and denitrification were higher in the bulk soil (Fig. 5) because the rhizosphere is nutrient depleted due to root uptake 30. This is indirectly confirmed by the fact that the rhizosphere affects nearly all these N cycle-related functions, whereby this depletion is alleviated by N fertilization (Fig. 5g-q). Nevertheless, cellulolysis, ureolysis and chitinolysis were more intensive in the rhizosphere, reflecting the increased abundance of bacteria degrading these substances. Members of Lysobacter (affiliated to Gammaproteobacteria) were abundant in the rhizosphere (Fig. 2) and are known to be chitinolytic bacteria 58. The methanol oxidation and methylotrophy genes are much more abundant in the rhizosphere than in the soil (except paddies). Methylotrophy is higher in the rhizosphere, but this difference is equalized in paddy soils because of the aerobic microenvironment around rice roots and because the available reduced C is strongly diluted in water excess.
In nature, specifically in the rhizosphere, plants are constantly challenged by thousands of microbial populations, including commensals, pathogens and symbionts. Plant pathogens and N-fixers (e.g. Rhizobium sp., etc.) are enriched in the rhizosphere (Fig. 5) because their reproduction and functioning are dependent on a plant host supply with organics. Although the rhizosphere is a fluctuating environment in which the microbiome rapidly evolves in space and time, the accumulating evidence verifies that plants shape their rhizosphere microbiome to their own benefit, making sophisticated use of the microbial functional repertoire 12. Nonetheless, it remains a challenge to clarify the equilibrium conditions for maintaining plant fitness, which involves establishing a balance between the passive attacks of pathogens and the active recruitment of beneficial bacteria.
By integrating sequencing data from several studies, we generalized the main differences in the microbiomes of rhizosphere and bulk soil regarding bacterial diversity, composition, selection of specific groups, co-occurrence network, and a very broad range of functions (Fig. 6). Bacterial diversity in the rhizosphere is reduced by 2.6-7% and represents a subset of the communities in bulk soil. The rhizosphere community composition is highly enriched with copiotrophic bacteria such as Proteobacteria and Bacteroidetes, while it is strongly depleted in Chloroflexi, Acidobacteria and Nitrospirae. As the rhizosphere has an organic C surplus and circulates nutrients quickly, the quicker-growing bacteria are overrepresented with functions related to lignocellulose degradation and plant pathogens, but depleted in the functions responsible for N cycling-related processes (except N fixation). The indirect proof of the generalizations presented here is that nitrogen fertilization alleviated nearly all rhizosphere effects on bacterial diversity and functions. This also confirms the common mineral N depletion against the background of excess C around roots. In conclusion, the selective effects of the rhizosphere in shaping microbial communities to some extent overrides the differences between soils, crops and climate. This makes the rhizosphere the strongest factor in forming the composition, structure and functions of the soil microbiome and, thus, a key factor in the cycling of biogenic elements.