New insights into functional divergence and adaptive evolution of uncultured bacteria in anammox community by complete genome-centric analysis

Abstract

Anaerobic ammonium-oxidation (anammox) bacteria play a crucial role in global nitrogen cycling and wastewater nitrogen removal, but they depend on mutualistic symbiosis with various other microorganisms for their survival. Although shotgun metagenomics based on short reads has been widely used in anammox research, metagenome-assembled genomes (MAGs) are often discontinuous and highly contaminated, which limits in-depth analyses of anammox communities. Here, for the first time, we performed Pacific Biosciences high-fidelity (HiFi) sequencing on the anammox granule sludge sample, and obtained 30 accurate and complete metagenome-assembled genomes (cMAGs). These cMAGs were obtained by selecting high-quality circular contigs from initial assemblies of long reads generated by HiFi sequencing, eliminating the need for Illumina short reads, binning, and reassembly. cMAG-centric analysis revealed divergences in general and nitrogen metabolism among members of the anammox community. Furthermore, we identified mobile genetic elements (MGEs) and putative horizontal gene transfer (HGT) events within these cMAGs to explore the adaptive evolution of the community. The results suggest that MGEs and HGT events, particularly transposons containing tnpA in anammox bacteria, might play important roles in the adaptive evolution of this anammox community. The cMAGs generated in the present study could be used to establish of a comprehensive database for anammox bacteria and associated microorganisms. Our findings highlight the advantages of HiFi sequencing for the studies of complex cultures such as anammox communities and advance our understanding of anammox communities.