Antibacterial efficacy of acetone extract, from different parts of sporophyte of the medicinal fern Cyclosorus interruptus (Willd.) H. Ito (Thelypteridaceae, Pteridophyta), was studied by disc diffusion method by using Staphylococcus aureus as test organism. Sporophytic parts such as rhizome, roots, rachis, sporangia and spores showed nil activity. The in vitro cultured gametophytes also showed nil activity. Only leafy parts of the sporophyte such as, croziers (young leaves), sterile and fertile mature leaves and rachis with leaves showed antibacterial activity with the inhibition zone from 18 to 28mm. The microscopical epidermal glands on leaves showed maximum antibacterial efficacy. The crude acetone extract and the ethyl acetate fraction of the crude extract of the epidermal glands showed comparatively higher degree of antibacterial efficacy followed by chloroform fraction of crude extract. All the six concentrations (5, 100, 250, 500, 750, 100μg) of chloroform fraction of crude extract showed antibacterial efficacy with the inhibition zone from 8-11mm. But the ethyl acetate fraction showed maximum size of inhibition zone 13mm in 100 μg. The acetone extracts of Cyclosorus interruptus revealed the highest antbacterial activity at a minimum concentration 31.255μg/100μl against S. aureus. Among the macroscopical and microscopical parts of sporophyte of the medicinal fern Cyclosorus interruptus, the leafy part of the sporophyte show maximum antibacterial activity for which the epidermal glands with various bioactive compounds play a major role.