Memory-bound <i>k</i> -mer selection for large and evolutionarily diverse reference libraries

Using <i>k</i>-mers to find sequence matches is increasingly used in many bioinformatic applications, including metagenomic sequence classification. The accuracy of these downstream applications relies on the density of the reference databases, which are rapidly growing. Although the increased density provides hope for improvements in accuracy, scalability is a concern. Reference <i>k</i>-mers are kept in the memory during the query time, and saving all <i>k</i>-mers of these ever-expanding databases is fast becoming impractical. Several strategies for subsampling have been proposed, including minimizers and finding taxon-specific <i>k</i>-mers. However, we contend that these strategies are inadequate, especially when reference sets are taxonomically imbalanced, as are most microbial libraries. In this paper, we explore approaches for selecting a fixed-size subset of <i>k</i>-mers present in an ultra-large data set to include in a library such that the classification of reads suffers the least. Our experiments demonstrate the limitations of existing approaches, especially for novel and poorly sampled groups. We propose a library construction algorithm called <i>k</i>-mer RANKer (KRANK) that combines several components, including a hierarchical selection strategy with adaptive size restrictions and an equitable coverage strategy. We implement KRANK in highly optimized code and combine it with the locality-sensitive hashing classifier CONSULT-II to build a taxonomic classification and profiling method. On several benchmarks, KRANK <i>k</i>-mer selection significantly reduces memory consumption with minimal loss in classification accuracy. We show in extensive analyses based on CAMI benchmarks that KRANK outperforms <i>k</i>-mer-based alternatives in terms of taxonomic profiling and comes close to the best marker-based methods in terms of accuracy.