Forty solvent fractions and 387 bacterial isolates of seven varieties of Korean domestic honey and manuka honey from New Zealand were screened for antimicrobial activity. The minimum inhibitory concentrations and minimum bactericidal concentrations of the honey fractions were determined; only Bacillus cereus ATCC 14579, ATCC 11778, and F4552 were inhibited by 11, 1, and 16, respectively, out of the 40 honey fractions. The bacterial isolates showed the highest incidence (30.2%) of antimicrobial activity against Listeria monocytogenes ATCC 15313. The growth of at least one of the five foodborne pathogens tested was inhibited by 109 of the 327 isolates (33.3%) from seven types of Korean domestic honey. The percentage of such isolates of manuka honey was significantly higher (76.7%). Solvent fractionation of honey could contribute to the detection of antimicrobial activity of the nonsugar compounds in honey. Moreover, the bacterial isolates from Korean domestic honey may be good sources for the natural antimicrobials used in the food industry and other related industries.