Leaf litter is an important energy source in forested streams, and is degraded primarily by fungi (mainly hyphomycetes) and bacteria. Not much is known about the diversity of non-hyhomycete/sporulating fungi, bacteria and actinomycetes on leaf litter and interactions among these groups. Three aspects of microbial ecology on decomposing leaves were examined. First, the diversity of fungi, bacteria and actinomycetes on decaying sugar maple and white oak leaves was examined for 181 days using denaturing gradient gel electrophoresis (DGGE). Fungi exhibited greater biomass and ribotype richness than bacteria. Temporal variation but no leaf-specific differences were observed in microbial community structure suggesting that the microorganisms on either leaf species were generalists and could exploit both resources equally well. Second, the community composition of fungi was examined using a traditional method (conidia staining) as well as cultivation and phylogenetic analysis. Conidia staining revealed that Tetrachaetum elegans was dominant, while 18S rRNA gene sequencing of cultured isolates revealed mainly non-hyphomycetes (Ascomycota) and a greater richness than microscopy. Only 12 isolates were conclusively identified of which two were hyphomycetes. Third, interactions between T. elegans and two bacterial isolates, CM8 and Kanr12 (antibiotic-resistant) studied under varying dissolved organic carbon (DOC) regimes showed no influence of DOC amendment on microbial biomass. No synergism/antagonism was observed between T. elegans and bacteria probably because nutrients were not limiting. Both bacteria belong to the metabolically versatile Cytophaga-Flavobacterium cluster, which may indicate that they could exploit the resources equally well. Hence, no discernible differences were observed in their response to the fungus. This study demonstrated that ecological niche of microorganisms, stage of decay and time of exposure are more important determinants of microbial community structure than leaf quality