Multidrug-resistant bacteria represent a growing concern and complex challenge in healthcare [1]. The presence of these microorganisms in diabetic foot ulcers (DFU) is responsible for high hospitalization and amputation rates [2]. Honey has demonstrated effectiveness in DFU treatment due to its anti-inflammatory and antibacterial properties, namely against bacterial biofilms and multidrug-resistant bacteria [3]. However, the variety of physical-chemical characteristics among different types of honey, such as the type of pollen, might confer different biological properties, potentially leading to differences in antimicrobial response [4].This study aimed to assess the effect of different types and concentrations of honey against bacterial biofilm. Seven different types of honey from the region of Trás-os-Montes (Portugal) were tested at three concentrations: i) 1xMinimum Inhibitory Concentration (MIC), ii) 5xMIC, and iii) 10xMIC, against biofilms of Staphylococcus aureus, Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Candida albicans. For Candida albicans, honey type-3, and type-4 removed 80.2% and 78.4% of biofilms, respectively, which was significantly higher than type-5 (8.87%), type-8 (27.5%), and type-9 (21.5%). Similarly, biofilm removal promoted by the honey type-2 (64.1%) and type-4 (57.2%) was greater than type-10 (28.5%) for E. coli. Principal Component Analysis suggested correlations between different pollen content and antimicrobial activity. Principal regression analyses were significant suggesting negative correlations of biofilm removal with pollen from Erica sp. and Sedum sp., and positive correlations with pollens from Corrigiola telephiifolia, and Jasione montana. Differences in bacterial responses may be due to variations in honey's pollen content and bacterial strain sensitivity.