To isolate and characterize keratinolytic fungi and bacteria from indigenous soils, a total of 80 samples were collected from Ghari Mori District. Khairpur, and these organisms were isolated using standard microbiological technique. The isolated keratinolytic microorganisms comprised: Absidia sp., Chrysosporium asperatum, Chrysosporium keratinophilum, Entomophthora coronata, Bacillus subtilis and Staphylococcus aureus and their keratinolytic properties were distinguished from the production of keratinase by measurement of zone of hydrolysis on skimmed milk agar (p<0.05). C.keratinophylum and B. subtilis produced largest zone among all the isolated species. The crude keratinase revealed that the optimum time for production of the enzyme was seven days, optimum temperature 30°C and optimum pH 9 for C.keratinophylum but for B. subtilis, the optimum time was three days, optimum temperature 37°C and optimum pH 7. The enzyme activity of C. keratinophylum and B. subtilis were determined to be 220 U/ml and 260 U/ml respectively (P<0.05).