Phospholipid fatty acids (PLFAs) were analysed at different time periods during composting of two waste mixtures rich in fats, M1 (22%) and M2 (39%), with the aim of monitoring changes in microbial community structure. The two mixtures consisted of a sludge sample collected from a vegetable oil refinery effluent treatment plant combined with household wastes. The PLFA profiles of both mixtures revealed that, at the start of the process, fungi and Gram-negative bacteria (G−) were more abundant in M2 than in M1. During the thermophilic phase, branched PLFA (i15:0, a15:0, i16:0, and i17:0) markers of Gram-positive bacteria (G+), became more abundant in M1, while G- bacteria were predominant in M2. The PLFA profiles in M1 representing non-specific, G+ and G− bacteria as well as fungi decreased during the cooling phase (maturation) while an increase was recorded in M2, which was richer in fats. The Shannon–Weaver diversity index (Ish) showed a greater increase during M1 composting (from 0.69 to 1.05), mainly for G+ bacteria and G− bacteria, than in M2 composting (from 0.79 to 0.84). Principal components and cluster analyses revealed a succession of different communities during composting, which varied from fungi and G− bacteria to G+ and thermophilic and thermotolerant G− bacteria. The end of composting was characterized by a reduction of all these microbial entities, especially for M1, except actinomycetes, which are associated with compost stability.