Microstructures of polycrystalline diamond thin films deposited on silicon (100) substrates which were pretreated by (a) wet polishing, (b) dry polishing, (c) sonication and (d) acid etching, respectively, have been studied by scanning electron microscopy (SEM) and high resolution transmission electron microscopy (HREM). The thin films were deposited by using a Hot Filament Assisted Chemical Vapor Deposition (HFCVD) technique, with a methane to hydrogen ratio of 0.5%. Atomic Force Microscopy (AFM) was used to characterize substrate surfaces after the different pretreatments. The wet and dry polishing produced substrates with scratch type of defects while the sonication and acid etching caused pit type of defects. Planar and cross-sectional SEM studies indicated that diamond films on the wet and dry polished surfaces were continuous and had a high density of nucleation sites while the films on the sonicated and acid etched surfaces were discontinuous in nature under the same deposition time (8 hours) and condition. Cross-sectional HREM studies did not reveal any intermediate layers such as SiC and amorphous carbon between the thin films and substrates. Tungsten carbide nano-sized crystals, however, were observed to be dispersed at the interface region. These particles were believed to be formed during carburizing a fresh tungsten filament and deposited on the substrate during initial stage of growth.