Ammonia-oxidizer community
Under nutrient cessation, the community structure of ammonia oxidizers (AOA, AOB, CladeA, and CladeB) was characterized by 454-pyrosequencing analysis. AOB OTUs increased monotonically after the cessation of N additions in the 30, 60, 90, and 120 kg N ha−1yr−1 treatments, whereas AOA, CladeA, and CladeB were unaffected (Fig. S1). Similar patterns emerged with community structure when PCA was examined (Fig. 2). Permutational analysis of variance revealed that plots that had previously been fertilized significantly influenced AOB community composition (F=3.60, P=0.002). AOA, CladeA, and CladeB did not differ among R0, R30, R60, R90, and R120.
Random matrix theory (RMT)-based molecular ecology networks (MENs) were constructed using OTU data to reveal the ecological interactions among four groups (AOA, AOB, CladeA, and CladeB) in soils (Fig. 3). The overall AOA, AOB, and comammox Nitrospira communities network showed distinct co-occurrence patterns (Fig. 3). Topological properties of MENs (Table S4) showed more nodes and links in the high fertilization treatments (455 nodes, 1636 links for R90, 426 nodes, 1604 links for R120) than the control (299 nodes, 1400 links). Most connections between OTUs were positive in the network. AOA had the most intensive associated with the networked communities in control, suggesting the greater ecological importance of AOA in constructing the overall network. In R30 and R60, although AOA built more connections than AOB and comammoxNitrospira groups, AOB trends to increase associations with the networked communities and greatly outnumbers AOAs in R90 and R120 soils.