Honey is known as an alternative remedy for the treatment of wounds. To evaluate the potential of five Hungarian honey types against wound-associated bacteria, in vitro microbiological assays were conducted on Pseudomonas aeruginosa, Staphylococcus epidermidis and methicillin-resistant Staphylococcus aureus (MRSA). Minimum inhibitory concentration (MIC) was determined with the broth macrodilution method, and biofilm degradation capacity was tested with a crystal violet assay. To understand the underlying mechanisms, the effects of honey treatments were assessed on bacterial membrane integrity and quorum sensing (QS). The highest antibacterial activity, indicated by the lowest MIC values, as well as the highest biofilm inhibition rates and membrane disruption, was displayed by chestnut and linden honeys. The most sensitive bacterium was S. epidermidis. Bacterial membrane degradation took place 40 min after treatment with honey solutions of at least a 40% concentration. Each honey sample exhibited anti-QS activity, which was most pronounced in the case of chestnut honey. It was concluded that the antibacterial, biofilm-inhibiting and anti-QS activities of linden and chestnut honeys were superior to those of acacia, goldenrod and milkweed honeys. In addition to the floral source, the antibacterial effect of honey is influenced by the microbial species treated. The use of honey in wound treatment can be justified by its diverse antibacterial mechanisms.