The Rivulidae fish family, which includes Neotropical seasonal killifishes, is one of the most diverse taxonomic groups in the aquatic systems of Caatinga in Brazil. Cynolebias and Hypsolebias genera, with 20 and 35 endemic species, respectively, concentrate the greatest diversity of rivulid species in the semiarid. Sixty-eight years after the first records of annual killifishes in the Mid-Northeastern Caatinga ecoregion (MNCE), only four valid species have been sampled in this area. Here we combined bibliographic surveys and recent samplings to investigate the distribution of seasonal rivulids in MNCE. Twenty-one records were obtained, nine of which are new localities, expanding the distribution of three species: Hypsolebias martinsi, H. antenori and Cynolebias microphthalmus. Hypsolebias longignatus is still only known from its type locality in Ceará, near the Environmental Protection Area in Pacoti River, and has not been sampled ever since its description in 2008. Among the four species present in MNCE, H. antenori is the only species occurring within the limits of a conservation unit in the Furna Feia National Park. Anthropogenic impacts were observed in most temporary habitats visited, which ranged from river channel to small ponds in cave entrances. All records are found in coastal basins that discharge in the northern coast of the MNCE, in Ceará and Rio Grande do Norte states, which are crucial for the conservation of the Caatinga’s killifishes. The results also evidenced the importance of karstic habitats in the Jandaíra Formation as potential biotopes for seasonal fish in MNCE. This information must be used to update the conservation status of these species and highlight the importance of strategies for preserving the Caatinga’s temporary aquatic habitats, which should be considered for environmental licensing purposes.

Few studies have focused on non-troglomorphic fishes occurring in Brazilian caves, especially those in the Caatinga region. The present study is the first survey of fishes from karstic areas of the Jandaíra Formation in Rio Grande do Norte State, northeastern Brazil. This region is characterized by a high concentration of caves and a rich subterranean biodiversity, especially of troglobitic invertebrates, but remains considered a gap on the knowledge of the subterranean ichthyofauna in Brazil. Four field expeditions were carried out covering two dry and two rainy seasons, in 2018 and 2019, in 23 localities in small river basins along the western part of the Jandaíra Formation. A total of 829 fish specimens, none of them troglomorphic, was captured and identified as belonging to 25 species of 12 families and five orders. Amongst them, four species are endemic of the Mid-Northeastern Caatinga ecoregion, and two non-native species were recorded. Habitats were classified into three categories: superficial, associated to cave, and cave. Although no troglomorphic fish species was found in this study, we recorded 64.1% of the Apodi-Mossoró river ichthyofauna occurring in caves or associated to caves, corroborating the hypothesis that part of the ichthyofauna exploits these environments as a refuge during the dry season. In addition, we suggest conservation policies for the maintenance of subterranean and aquatic semiarid ecosystems in the Jandaira formation, which is currently under threat due to anthropogenic activities, such as mining and deforestation.