The Temporal And Spatial Successions Of Eukaryotic Microbial Communities In Soybean Farmland Ecosystem

The plant rhizosphere,a narrow area that connects root and surrounding soil,harbors an extraordinary number of microbes.These microbes can contribute to plant growth,health,and nutrition.Recent advances in high-throughput sequencing have promoted the studies of rhizospheric microbial community.However,most rhizospheric microbiome studies mainly have focused on bacteria,leaving the eukaryotic microbes in rhizosphere seldom investigated.In the study,first,we investigated the biogeographic patterns and community diversities of fungi and protists in bulk soil and rhizosphere of soybean(Glycine max)collected from six ecological regions in China.We also combined the bacterial high-throughput data to compare the differences and similarities among co-occurrence patterns of bacteria,fungi and protists,and explored their interactions.Then we explored the assemblages and ecological statuses of abundant and rare eukaryotic microbial communities.Finally,we investigated the temporal successions of eukaryotic microbial communities in bulk soil and rhizosphere at different developmental stages of soybean.The compositions of the fungal and protist communities were investigated through MiSeq high-throughput sequencing of the internal transcribed spacer 1(ITS 1)region and V9 region of the 18 S rRNA gene.The main results are as follows:First,the community structures of eukaryotic microbes in rhizosphere differed from bulk soil significantly.Eukaryotic microbial ?-diversity was significantly lower in rhizosphere than in bulk soil.The number of depleted OTUs in rhizosphere was higher than enriched OTUs.Ascomycota dominated the fungal communities at phylum level in both bulk soil and rhizosphere.Comparing the two compartments,the relative abundances of Dothideomycetes and Eurotiomycetes were significantly higher in rhizosphere at class level,whereas the Agaricomycetes,Leotiomycetes and Sordariomycetes were significantly less abundant in rhizosphere.For fungal trophic mode,the relative abundances of pathotroph and pathotroph-saprotroph were significantly higer in rhizosphere than bulk soil,whereas saprotroph,symbiotroph and saprotroph-symbiotroph were significantly less abundant in rhizosphere than bulk soil.Rhizaria dominated the protist communities in both bulk soil and rhizosphere.Comparing the two compartments,the relative abundance of Rhizaria was significantly higher in rhizosphere,whereas the Apusozoa,Archaeplastida,Excavata,Hacrobia,and Stramenopiles were significantly less abundant in rhizosphere.For protist trophic mode,the relative abundance of bacterivores was significantly higher in rhizosphere than bulk soil,whereas phototrophs and saprotrophs were significantly less abundant in rhizosphere than bulk soil.Second,significant linear distance-decay relationships were observed between eukaryotic microbial community similarities and geographical distance in bulk soil and rhizosphere.And the distance-decay slopes were steeper in rhizosphere,indicating that historical processes(such as geographical separation)have a greater influence in rhizosphere.The results of permutational multivariate analysis of variance and unconstrained principal-coordinate analysis revealed that location represented the dominant source of variation.And compartment significantly impact the soybean root-associated fungal and protist communities.However,cultivar only significantly impact fungal communities of rhizosphere,and have no significant influence in protist communities of rhizosphere.The results of canonical correspondence analysis and variation partitioning revealed that distinct effects of geographical distance,edaphic and climatic factors on the assembly of eukaryotic microbial communities in bulk soil and rhizosphere.Third,the ecological networks clearly were unalike between bulk soil and rhizosphere.The networks were larger and more complex in bulk soil than in rhizosphere.The proportion of positive correlations in rhizosphere was higher than in bulk soil.The interactions mainly occurred between closely related species in taxonomy.We identified some keystone species based on network topological features of nodes,the OTUs belonging to Bryobacter,Massilia,Gemmatimonas,Acidibacter,Altererythrobacter and Reyranellaus were identified as keystone species of bacterial network in bulk soil;the OTUs belonging to Nocardioides,Caenimonas,Methylibium,Reyranella,Ramlibacter,Solirubrobacter,and Nonomuraea were identified as keystone species of bacterial network in rhizosphere.The OTUs belonging to Mortierella and Fusarium were identified as keystone species of fungal network in bulk soil;and the OTUs belonging to Paraphaeosphaeria,Mortierella and Microdochium were identified as keystone species of fungal network in rhizosphere.The OTUs belonging to bacterivores and phototrophs were identified as keystone species of protist network in bulk soil;and the OTUs belonging to omnivores and phototrophs were identified as keystone species of protist network in rhizosphere.The keystone species in the total network contained bacteria,fungi and protists,indicated the crucial ecological roles to play as integral parts of root-associated microbial co-occurrence networks in soybean fields.The proportions of negative correlations of protist-bacteria and protist-fungi interactions were higher in rhizosphere than in bulk soil,assuming more predator-prey relationships in rhizosphere.Fourth,distinct biogeographical patterns were observed between abundant and rare eukaryotic microbial communities in bulk soil and rhizosphere.The results of Sloan neutral model analysis revealed that stochastic processes played a more important role and deterministic processes played a less important role in shaping the rare taxa compared with abundant taxa.Co-occurrence network analysis showed that the importances of nodes were not depend on the relative abundances of them.No significant difference was observed in degree and betweenness centrality between abundant and rare taxa.Keystone species in networks contained abundant and rare taxa,suggested important roles of abundant and rare taxa in eukaryotic microbial networks.Finally,successions were observed in the eukaryotic microbial communities in bulk soil and rhizosphere during developmental stages of soybean.The succession rates were higher in rhizosphere than in bulk soil.The differences of eukaryotic microbial communities between bulk soil and rhizosphere increased with developmetal stages after branching stage,suggested that different directions of succeessions of the eukaryotic microbial communities in bulk soil and rhizosphere,and increasing selective power of plant roots.The results of permutational multivariate analysis of variance and unconstrained principal-coordinate analysis revealed that compartment had a greater impact on variation of eukaryotic microbial communities.Co-occurrence network analysis showed that different eukaryotic microbial co-occurrence networks were observed between bulk soil and rhizosphere at each developmental stage of soybean.And different co-occurrence patterns were observed in eukaryotic microbial communities at different developmental stages of soybean in each compartment.The largest-scale networks were observed in branching stage in bulk soil and rhizosphere.The network parameters in rhizosphere changed greater than in bulk soil during developmental stages of soybean.The compositions of keystone species of eukaryotic microbial networks were varied among different compartments and developmental stages.In conclusion,the geographical distributions and temporal successions were distinct between the eukaryotic microbial communities in bulk soil and rhizosphere.The importance of eukaryotic microbe was not depended on its abundance.Rare taxa also contributed to stability and ecological functions of eukaryotic microbial communities in bulk soil and rhizosphere.These results provide new views for the study of root-associated microbial communities.