In order to search for new antimicrobial agents, a number of 2-substituted 8-ethyl-5,8-dihydro-5-oxothiazolo[5,4-b]naphthyridine-6-carboxylic acid and their related compounds which consist of a new ring system were prepared. Reactions of the 6-amino-1-ethyl-7-mercapto-1,4-dihydro-4-oxonaphthyridine-3-carboxylic acid with acid, acid anhydride, acid chloride and ethylxanthate afforded several thiazolo[5,4-b]naphthyridine derivatives. Refluxing ethyl 7-chloro-1-ethyl-6-nitro-1,4-dihydro-4-oxonaphthyridine-3-carboxylate with KSCN in acetic acid gave directly the thiazole cyclization product. Reaction of 8-ethyl-2-methylthio-5,8-dihydro-5-oxothiazolo[5,4-b]naphthyridine-6-carboxylic acid with dimethyl sulfate gave various products, depending upon its reaction conditions. Some compounds obtained in this work exhibited high activities against gram-negative and gram-positive bacteria in vitro.