Vritra: gene-specific reference construction for species-resolved functional profiling of metagenomic and metatranscriptomic data
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Species-resolved profiling of microbial functional genes is important for linking microbial community composition to biological function, but existing functional profiling approaches are not designed to systematically define user-specified genes and resolve their contributing species. We developed Vritra ( V ersatile gene-guided R eads-identification with I mpartial T axonomic R efinement and A ssignment), a framework for constructing gene-specific reference databases for metagenomic and metatranscriptomic data. Vritra expands and refines the UniRef-based sequence space for user-specified target genes using sequence-similarity network connectivity and functional annotations, retains related homologs as decoys to reduce assignment ambiguity, and links the resulting sequences to standardized microbial taxonomy. Across genes involved in oxalate, urate, and bile acid metabolism, Vritra recovered >97% of sequences represented by established annotation resources for most evaluated genes while substantially expanding the represented sequence space. In real microbiome datasets, the expanded references increased recovery of target-gene reads by up to threefold for poorly annotated genes while maintaining high sequence identity. Application to population-based and publicly available microbiome datasets enabled species-resolved profiling of functional genes and revealed gene-specific associations with microbial taxa, dietary factors, and disease-related phenotypes. Vritra provides a scalable framework for translating continuously expanding sequence resources into gene-specific, species-resolved references for microbiome studies.
Importance
Microbial genes encode functions that can influence human health, yet it is often difficult to determine which microbial species carry out specific functions in complex microbiome communities. Existing approaches are well suited for broad functional profiling but provide limited flexibility for studying individual genes, particularly when those genes are poorly characterized or represented inconsistently across reference resources. Vritra addresses this gap by providing a framework for building gene-specific reference databases that connect functional genes with the microbial species that carry them. The approach can be applied to both metagenomic and metatranscriptomic data and can accommodate user-specified target genes, including genes from emerging or incompletely characterized pathways. By enabling species-resolved profiling of selected microbial functions, Vritra can help researchers investigate how specific microbial activities vary across individuals and relate to dietary, ecological, and disease-related factors. This framework provides a practical approach for studying microbial functions that may be overlooked by broad functional profiling methods.
