Anellovirus abundance as an indicator for viral metagenomic classifier utility in plasma samples.

Background: Viral metagenomics has expanded significantly in recent years due to advancements in next-generation sequencing, establishing it as the leading method for identifying emerging viruses. A crucial step in metagenomics is taxonomic classification, where sequence data is assigned to specific taxa, thereby enabling the characterization of species composition within a sample. Various taxonomic classifiers have been developed in recent years, each employing distinct classification approaches that produce varying results and abundance profiles, even when analyzing the same sample.

Methods: In this study, we propose using the identification of Torque Teno Viruses (TTVs), from the Anelloviridae family, as indicators to evaluate the performance of four short-read-based metagenomic classifiers: Kraken2, Kaiju, CLARK and DIAMOND, when evaluating human plasma samples.

Results: Our results show that each classifier assigns TTV species at different abundance levels, potentially influencing the interpretation of diversity within samples. Specifically, nucleotide-based classifiers tend to detect a broader range of TTV species, indicating higher sensitivity, while amino acid-based classifiers like DIAMOND and CLARK display lower abundance indices. Interestingly, despite employing different algorithms and data types (protein-based vs. nucleotide-based), Kaiju and Kraken2 performed similarly.

Conclusion: Our study underscores the critical impact of classifier selection on diversity indices in metagenomic analyses. Kaiju effectively assigned a wide variety of TTV species, demonstrating it did not require a high volume of reads to capture diversity. Nucleotide-based classifiers like CLARK and Kraken2 showed superior sensitivity, which is valuable for detecting emerging or rare viruses. At the same time, protein-based approaches such as DIAMOND and Kaiju proved robust for identifying known species with low variability.