Abstract The synthesis and microbiological evaluation of 7-, 8-and 9-nitro-1, 2, 4- trihalogenophenoxazin-3-one substrates with potential in the detection of nitroreductase- expressing pathogenic microorganisms are described. The 7-and 9- nitrotrihalogenophenoxazinone substrates were reduced by most Gram-negative microorganisms and were inhibitory to the growth of certain Gram-positive bacteria; ...