Abstract
Characterizing microbial strain diversity within complex mammalian gut microbiomes remains a formidable computational challenge due to extensive microheterogeneity, horizontal gene transfer, and structural repetitive regions. Conventional short-read metagenomic assemblers struggle to resolve micro-variants, while standard long-read assemblers frequently collapse distinct sub-species strains into single consensus contigs. Here, we present Panspecies-Assembler, an open-source, phase-aware computational framework specifically engineered for long-read Oxford Nanopore Technologies (ONT) sequencing data. Panspecies-Assembler integrates overlap-layout-consensus assembly graph construction with a novel variational Bayesian strain-phasing algorithm, enabling high-resolution de novo assembly and longitudinal tracking of individual bacterial strains within polymicrobial communities. Benchmarking Panspecies-Assembler against state-of-the-art metagenomic tools using both synthetic communities and human fecal metagenomes demonstrated a contig N50 of 2.8 Mb—a 4.2-fold enhancement over conventional assemblers—while successfully recovering 94.6% of low-abundance strain variants with over 99.8% consensus sequence accuracy. Furthermore, application to a longitudinal gut microbiome dataset enabled the tracking of strain-specific mobile genetic elements and micro-evolutionary shifts following dietary interventions. Panspecies-Assembler bridges a critical gap in metagenomic bioinformatics, offering an effective pipeline for high-resolution strain tracking in complex microbial ecosystems.
