Specimens of the palpimanid spider subfamily Otiothopinae Platnick are commonly collected in soil and leaf litter in tropical and subtropical environments (Platnick 1975; Cala-Riquelme et al. 2018). However, otiothopines are generally poorly represented in biological collections and usually collected in low numbers during field campaigns, with many species represented by only a single sex (Cala-Riquelme et al. 2018). These difficulties, combined with our scarce knowledge of the biology and natural history of these spiders, mean that there are regional gaps in our systematic understanding of the group. Thus, taxonomic additions and notes about natural history are essential in order to increase our knowledge of the group and gradually close those gaps. To date, ten species of the subfamily from Colombia are listed in the World Spider Catalog (2021), eight of which belong to the genus Otiothops MacLeay, 1839 and only two recently described in Fernandezina Birab?n, 1951. Colombian species of Fernandezina were described from two distant localities and different environments, with F. eduardoi Cala-Riquelme, Quijano-Cuervo & Sabogal-Gonz?les, 2018 widely distributed along the Caribbean coast, and F. andersoni Cala-Riquelme & Agnarsson, 2018 known only from the Andean department of Boyac?. Here, we described a new species of Fernandezina from the Amazonian region of Colombia in Caquet? department, and present new data for F. eduardoi. A distribution map for the Colombian species of Fernandezina is also included. The type specimens examined are deposited in the Instituto Alexander Von Humboldt (IAvH-I, J. C. Neita). The multifocal images of the copulatory structures and the measurements of the specimens were taken with a Leica MC-190 HD digital camera attached to an S8AP0 Leica stereomicroscope with extended focal range. Multifocal images were assembled using Helicon Focus (5.3.14). The general format of descriptions follows Platnick (1975). Measurements are given in millimeters. Coloration patterns were described based on specimens preserved in 70?80% ethanol.