Myxobacteria are fascinating micro-predators due to their extraordinary social lifestyle, which is unique in the bacterial domain. These taxa are metabolically active in the soil microbial food web and control populations of soil microbes. However, the effects of fertilisation treatments on predatory myxobacteria in agricultural systems are often overlooked. Here, the high-throughput absolute abundance quantification (HAAQ) method was employed to investigate the abundance and cell density of myxobacteria in the Red Soil Experimental Station fields following 29 years of fertilisation. Using 16S rRNA gene amplicons, we detected a total of 419 myxobacterial operational taxonomic units (OTUs), accounting for 0.25-2.70% of the total bacterial abundance. Significantly different myxobacterial communities were found between nitrogen-fertilised (N_cluster) and manure-fertilised (M_cluster) samples by principal coordinate analysis (PCoA), analysis of similarities (ANOSIM), and Manhattan analysis (p < 0.05). N fertiliser treatments significantly decreased the myxobacterial abundance and copy number, species accumulation index (S), and Shannon index (p < 0.05). Furthermore, UpSet plots showed that the OTU number in the N fertiliser treatment was only 24.4% of that in the M treatment, as the application of N decreased the number of low-abundance myxobacterial OTUs. In addition, network analysis, redundancy analysis (RDA), and random forest (RF) analysis showed that myxobacterial abundance and copy number were the most important variables predicting the soil bacterial community and functional gene α- and β-diversity (P < 0.05). Our findings imply that soil acidification caused by the application of nitrogen fertilisers is the most important driver of the decrease in the myxobacterial abundance and copy number in the soil. We suggest that the changes in the abundance and number of myxobacteria are strongly correlated with the overall bacterial α- and β-diversity indices. In addition, such changes may be an important factor in the overall changes in microbial communities.