Pangenome analysis reveals both niche-specific "specialists" and microbial "side hustlers" in colorectal cancer microbiomes.

The gut microbiome is reproducibly implicated in colorectal cancer (CRC), yet the inter-study and interpersonal variability of certain species associations suggests that CRC microbiomes may be defined by convergent functional states achievable by phylogenetically diverse organisms. Distinguishing lineage-conserved "specialists" from taxonomically diverse "side hustlers"-organisms whose shared functional traits are dispersed across phylogenetically distant lineages-offers complementary translational insights: "specialists" are primary candidates for lineage-targeted biomarkers and inhibitors, while the shared functional architecture of "side hustlers" may reveal high-priority potential therapeutic targets robust to inter-individual variability. Here, we quantify phylogenetic coherence (monophyly) of 3,711 co-associated gene bins (CAGs) across 13 bacterial species and evaluate CRC associations across three independent cohorts, identifying hundreds of CAGs associated with CRC or health across a spectrum of monophyly scores, indicating that both states harbor a mixture of "specialist" and "side hustler" gene content. Strikingly, in Faecalibacterium prausnitzii-a species with a complex relationship to CRC-health-associated CAGs exhibited significantly higher monophyly scores than CRC-associated CAGs, consistent with health-linked traits being lineage-conserved while CRC-linked traits behave as polyphyletically distributed, potentially mobile "side hustlers." Across multiple CRC-associated species, we observe functional convergence in gene bins encoding Type IV secretion systems (T4SS), TonB-dependent receptors, and RagB/SusD nutrient uptake proteins. Fusobacterium animalis strains encode T4SS elements across bins with variable phylogenetic origins, representing simultaneous "specialist" and "side hustler" strategies within a single species. Pairwise interaction analysis further reveals synergistic interspecies associations, including co-occurrence of F. animalis and Clostridium scindens gene bins associated with a CRC probability of > 90%, suggesting that microbial "side hustlers" may amplify oncogenic risk through ecological interactions invisible to species-level analysis. These findings provide proof-of-principle that an ecological and evolutionary lens on the CRC microbiome can identify shared functional vulnerabilities and lineage-specific targets for microbiome-based cancer prevention.