Negativeome characterization and decontamination in early-life virome studies

• Published in: Nature communications Vol. 16; no. 1; pp. 6190 - 12
• Main Authors: Kuzub, Nataliia, Kurilshikov, Alexander, Zhernakova, Alexandra, Garmaeva, Sanzhima
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 04.07.2025; Nature Publishing Group; Nature Portfolio
• Subjects: 45; 45/22; 45/23; 49; 631/114/129/2043; 631/326/596/2142; Age; Babies; Biomass; Contaminants; Contamination; Datasets; Decontamination; Decontamination - methods; Feces - virology; Female; Gastrointestinal Microbiome - genetics; Genomes; Gut microbiota; Humanities and Social Sciences; Humans; Infant; Infant, Newborn; Infants; Male; multidisciplinary; Science; Science (multidisciplinary); Taxonomy; Virome - genetics; Viruses; Viruses - classification; Viruses - genetics; Viruses - isolation & purification
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-025-61478-7

Contaminant sequences of external origin complicate the study of host-associated viromes, particularly in low-biomass samples obtained through viral-like particle (VLP) enrichment. However, the prevalence and impact of external contaminants on low-biomass samples are under-studied. Here, we analyze 1321 gut virome samples and 55 negative controls (NCs) from four early-life virome studies. Virus sequences identified in NCs, termed negativeome, were used as a proxy for the contamination assessment. We show that 61% of samples share at least one identical strain with negativeome, likely representing external contamination. While the median abundance of contaminant strains in these samples is only 1%, it ranges from 0 to 99% and exceeds 10% in 11% of infant samples. We further demonstrate that contamination is largely study-specific and has a greater impact on infant samples than on maternal samples. Based on our results, we propose a contamination assessment method using a publicly available database of sequences detected in NCs and a strain-level decontamination strategy. Contamination significantly impacts low-biomass studies. Here, the authors highlight its pervasive and study-specific effects, particularly on early-life virome samples, and offer practical strategies to mitigate contamination.