Microorganisms enter the flotation process mainly from intake water and ore material. The diversity and number of microorganisms can vary significantly from mine to mine. In flotation, the conditions including oxygen levels, temperature, and nutrients from ore, intake water, and reagents are often favorable for the microbial growth. The mining industry aims to close the water loops, which is expected to result in the accumulation of microorganisms in the process waters with potential effects on flotation performance. Bioflotation, bioleaching, and bio-oxidation have been studied for decades as tools for concentrating and dissolving minerals. In contrast, there is limited scientific literature or industrial knowledge about microorganisms that naturally inhabit and prevail in minerals processing applications over a wide pH range. Microorganisms affect minerals when they selectively attach to the surfaces, produce extracellular polymeric substances (EPS) and polysaccharides, oxidize or reduce the minerals, change the pH and Eh of the process solution, and degrade organic flotation chemicals. Microorganisms contain different structural components that affect their surface chemistry, charge, and behavior in flotation, but these properties may also change via adaptation and solution conditions. Almost all studies on flotation have focused on chemical and physical parameters, and the role of naturally occurring microorganisms has remained underexplored. Advances in genomics and proteomics offer possibilities to describe not only which microorganisms are present, but also which physiological functions are being exercised. This article reviews the current knowledge of microorganisms in various mineral processes, identifies potential microbe–mineral interactions in flotation, describes the gaps in current knowledge, and concludes with the potential effects of microorganisms on flotation, especially in closed water loops.