Bacterial FeAs2 mineralization was found in the reddish-brown microbial mats that have grown on the walls of the drainage systems of Masutomi Hot Springs, Yamanashi Prefecture, Japan. The reddish-brown microbial mats, which are mainly composed of bacilliform and coccoid types of bacteria, have been analyzed and observed by microtechniques to interpret the bacterial biomineralization and search for the clues to bioremediation. These bacteria accumulate Fe and As along with other trace elements to form various biominerals. The electron diffraction (ED) pattern of the bacterial capsule identified lollingite (FeAs2) and calcite (CaCO3) on the surface of the cell. Based on Fourier-transform infrared absorbance spectroscopy (FT–IR) analysis, the presence of organic components such as C—H, C=O, CNH, –COOH, and N—H in the reddish-brown microbial mats emphasized the metal-binding potential of the bacteria. X-ray diffraction (XRD) data showed the poorly crystalline character of the precipitates, which consist mainly of hydrous iron oxides (2.7 Å (1 Å = 0.1 nm)). The FeAs2 biominerals form by adsorption onto the bacterial cell wall, as demonstrated by microscopic observations and spectroscopic analysis. These showed that bacteria in the reddish-brown microbial mats have the ability to form biominerals with heavy metals and toxic metalloids like As. Particularly significant in hot spring environments is the role of symbiotic and toxic-resistant bacteria, which have the ability to adapt to high As concentrations. Bacterial FeAs2 mineralization might also be considered a mechanism by which toxic As is removed from the aquatic ecosystem. The results provide evidence for detoxification processes and offer clues to possible methods of bioremediation.