Bacterial non-isoprenoid alkyl glycerol ether lipids (AGEs) are widespread in the environment but little is known about their biological precursors and mode of formation as a function of varying growth condi- tions. Here, we searched for the presence of AGEs in five pure strains of mesophilic (growth temperature between 20 and 40 C) and heterotrophic sulfate reducing bacteria from the family Desulfobacteraceae grown on a wide range of hydrocarbons and organic acids as sole carbon and energy source. Gas chro- matography–mass spectrometry (GC–MS) analysis of hydrolyzed cells revealed the presence of mono- alkyl glycerols (MAGEs) in one Desulfatiferula species and of MAGEs and dialkyl glycerols (DAGEs) in two Desulfatibacillum species. Species from the genus Desulfococcus did not produce AGEs. The ether lipid composition appeared strongly dependent on the nature and the chain length of the carbon substrate, with few to > 50 homologues of AGEs formed from a single substrate. Growth on long chain n-alkyl com- pounds (C14 to C18 alk-1-enes and fatty acids) generated a much lower diversity of MAGEs and DAGEs than short chain substrates (nonanoate, octanoate and pyruvate) or isoprenoid alkenes (phytadienes). Such variation in alkyl glycerol production could be linked to the distinct pathways involved in the meta- bolism of the different substrates. Despite this substrate-dependent AGE composition, the average chain length and level of branching of each class of ether lipid remained remarkably stable whatever the growth substrate and the number of MAGE and DAGE homologues formed, indicating a compositional control of ether lipids by heterotrophic bacteria to maintain optimal membrane properties.