FHV infection and replicon expression modulate phospholipid levels in Drosophila S2 cells. (A) Total PC levels in S2 cells infected with FHV or treated with miltefosine (left graph) or expressing an FHV RNA1 replicon (right graph). Total PC levels were determined in cells 24 h after infection, treatment, or replicon induction and are expressed relative to total cellular protein levels. (B) Phospholipid species distribution in control S2 cells, cells infected with FHV, or cells treated with miltefosine or oleic acid. Relative phospholipid species content is expressed as a molar percentage of total phospholipid content and was determined by ESI-MS/MS. (C) PC acyl chain length in mock, FHV-infected, and oleic acid-treated S2 cells. Total PC chain length was determined by ESI-MS/MS and represents the total number of carbons from both acyl chains. (D) PC acyl chain saturation in mock, FHV-infected, and oleic acid-treated S2 cells. Total number of C = C double bonds in PC species was determined by ESI-MS/MS and represents the total number of double bonds in both acyl chains. The fraction of PC species with greater than two double bonds was 0.5 to 1.5% for both mock and FHV-infected cells (see Additional File 4), and therefore these results were not included in the graph. (E) PC, PE, and PI species with acyl chains containing a total of 34 carbons and 2 double bonds in mock and FHV-infected S2 cells. P-value < 0.05* or 0.005**.