<i>Odoribacter splanchnicus</i> lipooligosaccharide: an uncommon structure with weak immunostimulatory activity

<i>Odoribacter splanchnicus</i>, a prominent member of the human gut microbiota, has emerged as a promising next-generation probiotic due to its anti-inflammatory properties and association with intestinal health. While previous studies have demonstrated its ability to attenuate inflammation and protect epithelial integrity, the molecular basis underlying these effects remained elusive. In this study, we provide the first comprehensive structural and functional characterization of the <i>O. splanchnicus</i> lipooligosaccharide (LOS), a major immunomodulatory surface component. NMR spectroscopy and mass spectrometry revealed a heterogeneous LOS structure composed of a mono-phosphorylated tetra- and penta-acylated lipid A backbone and a highly branched, short core oligosaccharide featuring unique residues such as 4-lactylmannose and galactosaminuronic acid. Small-angle X-ray scattering demonstrated that the LOS self-assembles into core-shell spherical micelles, with a large oligosaccharide corona shielding the lipid A, thereby limiting its accessibility to the innate immunity TLR4/MD-2 receptor complex. Accordingly, the <i>O. splanchnicus</i> LOS exhibited minimal TLR4 activation and failed to induce IL-8 secretion in enterocyte cell lines, in stark contrast to proinflammatory <i>Escherichia coli</i> LPS; in addition, it potently antagonized <i>E. coli</i> LPS-induced inflammation. These findings identify the LOS as a key effector of the <i>O. splanchnicus</i> anti-inflammatory phenotype and support its further development as a live biotherapeutic agent.