M. Tech. (Department of Biotechnology, Faculty of Applied and Computer Science), Vaal University of Technology| / Pigeon droppings, found in abundance in most cities and towns where pigeons are found, are a source of potential yeast and molds into the environment. Invasive fungal infections are a cause of morbidity and often mortality in immunocompromised individuals. The objective of this study was to the identification of bacterial and mold agents from pigeon droppings. Pigeon droppings samples were collected from three locations during the winter and summer months and studied for the occurrence of bacteria, yeast and molds by utilising culture-independent techniques. Amplification of the 16S rDNA gene and the internal transcribed spacer (ITS) region, cloning and ARDRA and DGGE were used for the characterisation of the microbial populations followed by sequencing. Several mold and yeasts, as well as bacteria were found to be present in pigeon droppings, which can spread into the environment and be transmitted to immunocompromised individuals and children. DGGE analysis of the bacterial communities revealed banding patterns that clustered all but one winter samples and all summer samples, showing a high similarity among the microbial members in both seasons and sample locations. Fungal DGGE analysis revealed clusters that grouped summer and winter samples from Johannesburg and Pretoria while VUT samples were clustered on their own. From the identification of fungal and bacterial DNA, Cryptococcus species was the majority of fungi isolated from the dropping samples. Geotrichum, Kazachstania and Fusarium species were isolated from phylotypes obtained from ITS amplicons analysed by ARDRA. Lactobacillus and Enteroccoccus species, organisms usually found in the gastrointestinal tract were the common bacterial members identified. The results showed no difference in microbial communities across all sample locations, while seasonal changes also had no impact in microbial community patterns.