The genus Humidophila is mainly characterized by small cells, striae often composed by one elongated areola occluded by thin hymenes. Species are distributed globally and are common in moist sub-aerial habitats. We identified five Humidophila species found in the well-conserved freshwater bodies and subaerial wet habitats from the mountain region "Mananciais da Serra". Four previously known species (Humidophila brekkaensis, H. contenta, H. nienta, H. paracontenta) and one new species (Humidophila piraquarae sp. nov.) were described and illustrated based on light and scanning electron microscopy observations. Humidophila piraquarae sp. nov. is similar to H. nienta (Carter) Lowe, Kociolek, Johansen, Van de Vijver, Lange-Bertalot et Kopalová and H. undulata Lowe, Kociolek, Johansen, Van de Vijver, Lange-Bertalot et Kopalová, but is best distinguished by their prominent constriction, apices broadly rostrate-rounded, striae density, raised siliceous thickening in the apical region of the axial area, and raphe proximal endings laterally expanded in anchorshaped depressions. Addtionally, a list of Humidophila taxa already registered in Brazil is presented.