Composite Metagenome-Assembled Genomes Reduce the Quality of Public Genome Repositories

In their recent study, Espinoza et al. employ genome-resolved metagenomics to investigate supragingival plaque metagenomes of 88 individuals (1). The 34 metagenome-assembled genomes (MAGs) that the authors report include those that resolve to clades that have largely evaded cultivation efforts, such as Gracilibacteria (formerly GN02) and Saccharibacteria (formerly TM7) of the recently described Candidate Phyla Radiation (2). Generating new genomic insights into the understudied members of the human oral cavity is of critical importance for a comprehensive understanding of the microbial ecology and functioning of this biome, and we acknowledge the contribution of the authors on this front. However, the redundant occurrence of bacterial single-copy core genes suggests that more than half of the MAGs that Espinoza et al. report are composite genomes that do not meet the recent quality guidelines suggested by the community (3). Composite genomes that aggregate sequences originating from multiple distinct populations can yield misleading insights when treated and reported as single genomes (4).

To briefly demonstrate their composite nature, we refined some of the key Espinoza et al. MAGs through a previously described approach (5) and the data that the authors kindly provided (1). We found that MAG IV.A, MAG IV.B, and MAG III.A described multiple discrete populations with distinct distribution patterns across individuals (Fig. 1). A phylogenomic analysis of refined MAG IV.A genomes resolved to the candidate phylum Absconditabacteria (formerly SR1) and not to Gracilibacteria as reported by Espinoza et al. (Fig. 1D). A pangenomic analysis of the original and refined MAG III.A genomes with other publicly available Saccharibacteria genomes showed a 7-fold increase in the number of single-copy core genes (Fig. 1E). These findings demonstrate the potential implications of composite MAGs in comparative genomics studies where single-copy core genes are commonly used to infer diversity, phylogeny, and taxonomy (6). Composite MAGs can also lead to inaccurate ecological insights through inflated abundance and prevalence estimates. For instance, the original MAG III.A recruited a total of 1,849,593 reads from Espinoza et al. metagenomes; however, the most abundant refined III.A genome (MAG III.A.2, Fig. 1C) recruited only 629,291 reads.

Co-assembly of a large number of metagenomes that contain very closely related populations often hinders confident assignments of shared contigs into individual bins. Nevertheless, even when proper refinement is not possible, reporting composite MAGs as single genomes should be avoided. As of today, highly composite Espinoza et al. MAGs (Fig. 1 in this letter and Table 1 in the work of Espinoza et al.) are available as single genomes in public databases of the National Center for Biotechnology Information (NCBI).

The rapidly increasing number of MAGs in public databases already competes with the total number of microbial isolate genomes (3), and increasingly frequent studies that report large collections of MAGs offer a glimpse of the future (7–9). Despite their growing availability, metagenomes are inherently complex and demand researchers to orchestrate an intricate combination of rapidly evolving computational tools and approaches with many alternatives to reconstruct, characterize, and finalize MAGs. We must continue to champion studies such as the one by Espinoza et al. for their contribution to our collective effort to shed light on the darker branches of the ever-growing Tree of Life. At the same time, editors and reviewers of genome-resolved metagenomics studies should properly scrutinize the quality and accuracy of MAGs prior to their publication. A systematic failure at this will reduce the quality of public genome repositories while yielding adverse effects such as misleading insights into novel microbial groups and reduced trust among scientists in findings that emerge from genome-resolved metagenomics.