A Sphingomonadales-associated phenylalanine catabolic pathway is implicated in ARDS recovery through the modulation of alveolar macrophages

<title>Abstract</title> <p> <bold>Background</bold> Acute respiratory distress syndrome (ARDS) carries a high mortality despite supportive care, and broad host-targeted therapies have failed to improve survival. Emerging evidence suggests that respiratory microbiota and their metabolic activities may modulate ARDS pathogenesis, but the underlying mechanisms remain unclear. <bold>Results</bold> ARDS survivors demonstrated significantly greater beta-diversity and enrichment of the order <italic>Sphingomonadales</italic> relative to non-survivors, a finding successfully validated in the external Kitsios cohort. Functional profiling identified phenylalanine metabolism as the key discriminatory pathway, with phenylalanine 4-monooxygenase (PhhA; EC 1.14.16.1) significantly enriched in survivors and mild ARDS. <italic>Sphingomonadales</italic> abundance correlated inversely with sputum L-phenylalanine levels and whole-genome sequencing confirmed a conserved <italic>phhA</italic> module in <italic>N. resinovorum</italic> . In the murine injury model, intratracheal <italic>N. resinovorum</italic> significantly reduced lung injury indicators. Critically, heterologous expression of <italic>phhA</italic> in a non-pathogenic <italic>E. coli</italic> chassis fully recapitulated the protective effects of native <italic>N. resinovorum</italic> , establishing PhhA as both necessary and sufficient for microbial phenylalanine catabolism-mediated lung protection. Mechanistically, we identified alveolar macrophages (AMs) as the cellular target through which this protective axis operates, demonstrating that <italic>N. resinovorum</italic> selectively preserves the AM compartment and that AM depletion largely abolishes its protective effects. <bold>Conclusion</bold> These findings establish a functional <italic>Sphingomonadales</italic> -PhhA-phenylalanine catabolic axis in ARDS. Respiratory <italic>Sphingomonadales</italic> abundance correlates with favorable outcomes and enhanced phenylalanine-catabolic potential. <italic>N. resinovorum</italic> degrades local phenylalanine via PhhA, and heterologous <italic>phhA</italic> expression alone is sufficient to recapitulate lung protection. This protection requires alveolar macrophages, as their depletion abolishes the effect. Thus, microbial phenylalanine catabolism acts as a spatially localized enzymatic buffer that compensates for deficient host metabolism in the injured lung, representing a mechanistically defined target for microbiota-informed ARDS therapy. <bold>Trial registration</bold> ClinicalTrials.gov, NCT07380997. Registered January 24, 2026. </p>