Successful fecal microbiota transplants in post-antibiotic treated recurrent <i>Clostridioides difficile</i> patients induce acylcarnitine and sphingolipid lipidomic shifts

<i>Clostridioides difficile</i> infection (CDI) is an urgent public health threat in the United States, causing over 500,000 cases, 29,000 deaths, and $4.8 billion in healthcare costs annually. Although fecal microbiota transplants (FMTs) are more effective than antibiotics for recurrent CDI (rCDI), their risks highlight the need for regulated alternatives such as live biotherapeutic products (LBPs). The development of effective LBPs requires understanding the biological mechanisms underlying FMT success. Building on our prior metabolomic study identifying lipids as key factors in FMT efficacy, we used liquid chromatography, ion mobility spectrometry, collision-induced dissociation, and mass spectrometry (LC-IMS-CID-MS) to profile >850 lipid species across 26 classes. In stool samples from 15 rCDI patients collected pre- and post-FMT (2 weeks, 2 months, and 6 months), 397 lipids were confidently identified. FMT significantly altered the lipidome (adonis R² = 0.12, <i>p</i> < 0.001), with 96 lipid species across 18 classes showing differential abundance (mixed effects modeling, BH correction, <i>p</i> < 0.05). Medium and long-chain acylcarnitines decreased post-FMT, while very long-chain acylcarnitines and trihydroxy ceramides increased. Sphingolipids were elevated pre-FMT. Medium and long-chain acylcarnitines correlated with Enterobacteriaceae, known to utilize carnitine. These results implicate lipid-microbiota interactions in rCDI susceptibility and recovery, offering targets for next-generation LBPs.