The adsorption of microorganisms on the mineral surface is the base of microorganisms that are considered as mineral processing reagents. The principles of the use of a highly hydrophobic and negatively charged bacterium, Mycobacterium phlei, as a floc-culating-flotating agent for finely divided hematite were investigated. The flocculating-floating recovery is strongly dependent on the pH and the dosage of the bacterium. Generally the pH should be controlled over the range of 5.5-7, and the dosage should be controlled about 16 mg/L. The infrared spectrometry analysis indicates that the six functional groups of M. phlei, substituted aromatic compound groups, -(CH2)n-groups, -CH2(-CH3)groups, carbonyl groups, aromatic hydrocarbon groups, and carboxyl groups, are on the hematite surface, among which the first five ones contribute physical adsorption and only the carboxyl groups provide chemisorption. Microscopic analysis reveals that the dimensions and tight aggregation degree of the floes of hematite particles formed by M. phlei are also impacted by the pH and the content of M. phlei in flotation.