Plant development strongly influences the assembly of plant microbiomes
Uncovering the ecological principles and processes that underpin plant microbiome assembly and developmental dynamics is essential to advance fundamental understanding of co-evolution and future application of the crop microbiome to sustainable increase in farm productivity [9, 10, 70]. Our results demonstrate that maize microbiome assembly is mainly influenced by compartment niche and developmental stage regardless of farming regions and fertilization regimes. Further, plant microbiomes are more sensitive to plant developmental stages than soil microbiomes in terms of multiple microbial attributes (i.e. alpha diversity, community structure, assembly processes and interkingdom networks). These findings are consistent with previous studies showing that plant compartment is a determining factor shaping the assembly of plant-associated microbiomes [26-29, 42], and that plant seasonal status has significant effects on microbiomes in grasses phyllosphere and Arabidopsis rhizosphere [32, 71]. Further, metagenomic analysis in our study revealed that the functional diversity and enriched functional traits in the maize phylloplane varied across three stages. There was a stronger depletion effect in the maize phylloplane in comparison to fake plant phylloplane, and functional attributes involved in methyl-accepting chemotaxis and defense mechanisms were significantly enriched in the maize phylloplane. Together these results indicate that the plant host exerts a strong selection effect to recruit and filter specific microbial taxa and functions from nearby species pool during plant development [18, 72, 73]. Complementary to the previous finding that host selection via plant compartment niche and host genetics plays a dominant role in shaping plant microbiomes assembly [20, 26, 29, 31, 42], this work provides novel evidence that plant developmental stages profoundly influence not only plant microbiome assembly but also their functions.
The effects of plant developmental stage represented the dynamic effects of plant metabolism, exudation and immune-associated traits [9, 13, 74], and plant-associated microbes have strong chemotaxis activities towards plant signal molecules such as organic acids and sugars [34, 71, 75-77]. For instance, a recent work revealed that wheat root-released organic carbon varied dramatically across wheat growth stages and correlated with different microbial taxa [34]. It was also suggested that plant exudates and volatiles like coumarins, benzoxazinoids and triterpenes play key roles in shaping plant microbiomes during host development [17, 72, 76]. The effect of plant developmental stages on microbiome in this study included the effects from season-dependent environmental factors like air, dust and climate. Our results showed that both maize and fake plant phylloplane microbiomes had similar temporal patterns and shared more than one third of ZOTUs at each stage. Further, fake plant phylloplane microbiome contributed an increasing proportion as the source of the maize phylloplane microbiome over the time, indicating a part contribution of environmental temporal factors to crop microbiome assembly. These results presented strong field evidence showing that local air, dust and rainwater are the main sources of crop microbiome in phyllosphere. These findings significantly advance our knowledge on the source, driving force and potential function of phyllosphere microbiome, and further corroborated that the phylloplane acts as an important interface between the host, microbes, and the environment [30, 78-80]. Our results also showed that plant developmental stage had significant effects on the rhizosphere and bulk soil microbiomes, though it was much weaker than the site effects, implying that plants also have profound influence on soil microbiomes via strong rhizosphere effect [6, 35, 74]. Collectively, by examining the temporal dynamics of bacterial and fungal microbiomes in the soil-plant continuum of maize and fake plants phylloplane in geographically distant sites, this study considerably expanded our knowledge on the succession of plant microbiomes and their potential function under different temporal and spatial scales in field.
The differentiation in ecological roles of bacterial and fungal microbiomes across plant developmental stages
Bacteria and fungi have coevolved with their host for more than 400 million years and greatly contribute to numerous aspects of plant health and productivity [1, 8, 41]. In this study, bacterial-fungal interkingdom interaction patterns distinctly shifted across three developmental stages. Bacterial community possessed higher alpha diversity and network connectivity at the seedling stage while fungal diversity was higher at the mature stage. Moreover, bacterial and fungal taxa dominated network hubs at the seedling stage and the mature stage, respectively. These suggested that the host can selectively modulate microbial interactions to meet its requirement during plant development, as microbial network hubs were supposed to play crucial roles in maintaining plant health and nutrient [37, 40]. In addition, bacterial taxa at the seedling stage were better predictors of crop yield while fungal taxa at the mature stage did so. Metagenomic analysis further corroborated that maize phylloplane microbiome possessed higher functional diversity at the seedling stage than the other two stages. Importantly, more abundant genes associated with disease resistance were enriched at the seedling stage while C degradation and P transportation related genes were enriched at two late stages, which further support the ability of plants to manipulate microbial assembly as per physiological requirements [1, 73]. Based on the limited knowledge on the plant microbiome, it has been proposed that the dynamics of plant microbiome composition are a reflection of the current needs of the host plant [3, 40, 73], and represent the consequence of subtle changes in microbial selection strategy exerted by the host during plant development [1, 13, 73, 81]. Our results therefore support that bacteria may take a more important ecological role in the plant microbiome and host performance at the early stage, while fungi do so at the late stage. This finding is supported by the Null model analysis, which demonstrated the dominant effect of determinism on bacterial community and of stochasticity on fungal community at the seedling stage, but a reverse pattern at the mature stage.
We further found that the negative edges representing bacterial-fungal interkingdom correlations in network increased over the time, implying an increasing competition relationship between bacteria and fungi along plant development stages. It was suggested that microbial competitive interaction could positively influence microbiome stability [40, 82, 83]. Our study provides more empirical evidence on this and further support the argument that the host may facilitate host fitness and plant-microbiome balance by deterministic host selection during plant development. These findings provide new insights into complex interactions among the plant, microbes and the environment and provide essential information for the future development of tools to manipulate crop microbiome.
Keystone bacterial and fungal taxa and their ecological functions at different developmental stages
Our results suggest that the composition and potential functions of plant microbiomes change across plant growth and bacterial phyla Actinobacteria and Bacteroidetes were more sensitive in response to plant developmental stage. More abundant Actinobacteria were observed at the seedling stage than at two late stages in plant compartments. Actinobacteria are well known as antagonistic bacteria excreting antibiotic compounds that provide protection against plant pathogens [84-86]. For example, a recent work showed that the enrichment of protective microbes like Actinobacteria in the rhizosphere could facilitate disease suppression [85]. Furthermore, some ZOTUs within families Burkholderiaceae, Streptomycetaceae and Rhizobiaceae were significantly enriched in plant compartment niches at the seedling stage. The members within Burkholderiaceae and Rhizobiaceae are important diazotrophs and plant growth promoting bacteria (PGPR) [1, 6, 12], and the members within Streptomycetaceae are well-known antibiotic-producing bacteria [41, 87, 88]. Coincidently, metagenomic analysis for the maize phylloplane suggested that functional genes involved in disease resistance and N2 fixation were significantly enriched at the seedling stage. In addition, bacterial communities in the rhizosphere and bulk soils showed significant correlations with nitrogenase activity across three developmental stages, and the bacterial functional group “nitrite respiration” was identified as the network hubs at the seedling stage. These findings provide evidence for the ecological importance of crop bacterial microbiomes in maintaining plant health and nutrient requirement at the early stage.
For fungal community, classes Sordariomycetes and Dothideomycetes were more sensitive to plant development, with Dothideomycetes increasing over the time in the rhizoplane but decreasing in the root endosphere. Previous works have shown that Sordariomycetes and Dothideomycetes are the most dominant fungal taxa in soils and plant compartments, respectively, and that class Dothideomycetes comprises a highly diverse range of fungi including endophytes, epiphytes and plant pathogens [42, 89]. In addition, many members within Dothideomycetes are also identified as saprotrophic fungi functioning in wood and leaf-litter decomposition and nutrient cycling [89, 90]. Notably, Dothideomycetes predominated phylloplane mycobiomes of both fake plant and maize in two distant study sites across three developmental stages. It was suggested that Dothideomycetes are the dominant fugal taxa of air microbiomes [90]. This indicated that Dothideomycetes in fake plant and maize phylloplanes might be mainly dispersed from air. The reverse dynamics of Dothideomycetes in the rhizoplane and root endosphere indicated multiple ecological roles of Dothideomycetes during host growth, however, its biological functions need further exploration. Furthermore, some fungal ZOTUs affiliating within families Coniothyriaceae, Mycosphaerellaceae and Symmetrosporaceae were identified as network hubs and significantly enriched in plant compartments at the mature stage. Some members of families Coniothyriaceae and Mycosphaerellaceae within Dothideomycetes are important saprobes with cellulose- and carbohydrate-degrading ability [90, 91]. Coincidently, we found that most network hubs in both taxonomic and functional networks of the mature stage belonged to fungal functional group “Saprotroph”. Moreover, fungal communities in the rhizosphere and bulk soils had significant correlations with C cycling related enzymes like β-glucosidase across three developmental stages. These results suggest that fungal taxa play key roles in regulating plant C cycles like decomposition of plant residues at the late stage. This indicates that crop mycobiomes may play an increasing ecological role as the decomposers with the aging of the plant, and the host plant may be passively occupied by saprophytic fungi as the consequence of reduced host immunity.
Collectively, our study demonstrates that plant is able to recruit specific microbial taxa with desire functions to meet their health and nutrient requirements at different developmental stages. However, the molecular mechanisms governing plant-microbiome interactions during host development and the ecological and biological functions of crop microbiomes in facing climate challenge and achieving sustainable agriculture are not fully understood and need further exploration [10, 92, 93].