Antimicrobial drugs used for prophylactic or therapeutic purposes in human, veterinary and agricultural purposes were favoring the survival and spread of resistant organisms[19]. The appearances of multidrug resistant pathogenic strains caused substantial morbidity and mortality especially among the elderly and immuno-compromised patients. To overcome this situation, there is an interest to improve or discover novel class antibiotics that have different mechanisms of action worldwide[20],[21]. The continuous screening of secondary microbial products produced from potential bacterial taxa was important to discover novel chemicals for the development of new therapeutic agents[22]. Recently, many scientists are searching new antibiotics from different untouched habitats to find out for their productions of antibiotics[23]. In Ethiopia, no significant studies have been conducted so far to isolate and evaluate actinomycetes from different freshwater habitats that could produce useful antibiotics. Therefore, present study is intended to isolate, screen and characterize antibiotic producing actinomycetes from water and sediments of Lake Tana, Ethiopia.