We have combined mineralogy, organic geochemistry and molecular microbiology to study hydrothermal deposits from Solfatara Crater, a geologically young volcanic formation (∼4,000 years old) displaying hot (45–95°C) and acidic (pH 1.7) mud pools and fumaroles. The search for inorganic (mineral) biosignatures revealed the presence of delicate structures, most likely mineralized extracellular polymers (EPSs), and the presence of potential biologically induced minerals: sulfides, sulfates (barite and alunite), elemental sulfur, and iron oxides. Geochemical analyses revealed a low total organic carbon content, 0.13 to 0.53%, displaying δ13C values from −17.09 to −27.39‰, and total nitrogen contents from 0.03 to 0.12%, which are characteristic of hydrothermal systems and suggest the presence of autotrophic carbon fixation. Lipid biomarker analysis showed the presence of hopanoids and linear alkanes, and the absence of detectable steroids, implying the occurrence of bacteria in our samples. We constructed 16S rRNA gene libraries from the environmental samples. Most environmental sequences obtained were affiliated to the Alpha- and Betaproteobacteria (Hydrogenophilus-like), the Acidobacteria, and to a lesser extent, the Gammaproteobacteria and Actinobacteria. When known, the closest cultivated relatives were often thermophilic or thermotolerant bacteria oxidizing iron, hydrogen, or methane/methanol, suggesting an important microbial contribution to the formation of biominerals.