Ratio of carbon and nitrogen in fertilizer treatment drives distinct rhizosphere microbial community composition and co-occurrence networks

Fertilization is the main strategy to accelerate vegetation restoration and improve the rhizosphere microbial community in the northeast China. However, the responses of rhizosphere microbial community structure, specific microbial community and symbiotic pattern to manure fertilization in grassland (alfalfa only) are not well clear. In this study, the variation of bacterial community structures in R_Manure (extracted liquid of fermented cow manure), E_Manure (extracted residue of fermented cow manure), F_Manure (full fermented cow manure), and Control (without fermented cow manure) collected from the rhizosphere microbial community of alfalfa were analyzed by the application of an Illumina HiSeq high-throughput sequencing technique. A total of 62,862 microbial operational taxonomic units (OTUs) were detected and derived from 21 phyla of known bacteria. The dominant bacteria in the rhizosphere include Proteobacteria (70.20%), Acidobacteria (1.24%), Actinobacteria (2.11%), Bacteroidetes (6.15%), Firmicutes (4.21%), and Chlorofexi (2.13%) accounting for 86% of the dominant phyla in all treatments. At the genus level, the dominant genus include NB1-j, Lysobacter, Alphaproteobacteria, Subgroup_6, Actinomarinales, Saccharimonadales, Aneurinibacillus, MO-CFX2, SBR1031, Caldilineaceae, and so on with the average relative abundance (RA) of 1.76%, 1.52%, 1.30%, 1.24%, 1.61%, 2.39%, 1.36%, 1.42%, 1.27%, and 1.03%, respectively. Bacterial diversities and community structures were significantly differentiated by different treatments of fertilization. The results of community structure composition showed that R_Manure treatment significantly increased the population abundance of Firmicutes, Chlorofexi, and Patescibacteria by 34.32%, 6.85%, and 2.70%, and decreased the population abundance of Proteobacteria and Actinobacteria by 16.83% and 1.04%, respectively. In addition, it showed that all treatments significantly resulted in an increase or decrease at the genus level. R_Manure had the higher richness and diversity of the bacterial community, with the greatest topology attributes of the co-occurrence networks. Through the analysis of the molecular ecological network (MENA), the co-occurrence networks had a shorter average path distance and diameter in R_Manure than in others, implying more stability to environmental changes. Redundancy analysis (RDA) showed that the ratio of carbon and nitrogen (C/N) was the main factor affecting rhizosphere microbial community composition while driving distinct rhizosphere bacterial community and its co-occurrence networks. The R_Manure associated with more C/N had relatively complex microbial co-occurrence network with a large number of nodes and edges, while the microbial network of others associated with less C/N had fewer taxa with loose mutual interactions. These results suggested that organic fertilizer with high C/N can regulate the rhizosphere microorganism, while high C/N can determine bacterial community structures, specific bacterial taxa, and their relationships with the nodule size of alfalfa. These significant changes can be used to evaluate soil fertility and fertilizer management in the artificial grassland system, while the potential biological indicators of the rhizosphere microbial community will play an important role in future eco-agriculture.