The present work was aimed in the Semmalai hills of Pudukkottai district to assess the density and diversity of fungal flora. To isolate and identify the fungi from the soil along with some physicochemical parameters like pH, electrical conductivity organic carbon and nitrogen etc., the month of January to December 2012. It has been estimated that 1.5 million fungal species are present in natural ecosystem, but only 5-10% have been described formally. During the study, a diversity of fungal strains were isolated and identified from the soil sample. Thus, the study on fungal diversity provides a basis for estimating the important functional role of fungi in soil ecosystem. The most common genera isolated from the soil samples included Aspergillus, Alternaria, Bipolaris and Fusarium.