Persistent high-risk human papillomavirus (HPV) infection is associated with the cervicovaginal microbiome, yet the mechanisms linking bacterial metabolism to viral persistence remain unclear. This study investigated whether lipid metabolites produced by vaginal bacteria influence HPV18 oncogene expression using an episomal HPV18 keratinocyte model (NIKS18). Conditioned media (CM) were generated from Lactobacillus crispatus DSM13335 (LC), Lactobacillus iners DSM20584 (LI), Gardnerella vaginalis DSM4944 (GV), and Prevotella bivia 12C1 (PB) cultured anaerobically in Vaginal Microbe Media v2 (VMM2). Lipidomic profiling by LC-MS (LPOS/LNEG) characterised bacterial metabolites, and NIKS18 cells were treated for 24 h with 2.5% or 5% CM before E6/E7 transcript quantification by RT–qPCR using REST and ΔΔCt analysis. GV CM displayed the highest lipid variability (median CV 30.5%) and enriched long-chain PUFAs (DHA 22:6, fold change 2.1, p<0.01) while significantly depleting lysophospholipids (LPE/LPA/LPI classes, fold change 0.55–0.70, p<0.05). PB CM significantly increased lysophosphatidylethanolamines (fold change 2.4–3.1), and LC CM increased sphingomyelin/ceramide classes (fold change 1.3–1.7, p<0.05). Vehicle control (VMM2) alone upregulated E6/E7 compared with untreated cells, highlighting lipid effects of the medium. When normalised to VMM2, LC and PB CM at 5% significantly upregulated E6/E7 expression (~1.5-fold, p<0.05, REST), whereas 2.5% CM caused minor decreases. To our knowledge, this is the first LC–MS lipidomic characterisation of cervicovaginal bacterial conditioned media (CM) benchmarked against a matched vehicle control. Distinct bacterial lipid signatures corresponded with specific HPV18 transcriptional responses, suggesting cervicovaginal microbial lipids may modulate viral persistence, potentially via membrane-associated PKC/YAP signalling.