Pareiorhaphis hystrix is a widely distributed species, occurring in the upper and middle Uruguay River and in the Taquari River basin, Patos Lagoon system, southern Brazil. Morphological variation has been detected throughout the distribution of P. hystrix, and this work seeks to test the conspecific nature of populations in several occurrence areas. Specimens from six areas in the Uruguay River basin and three in the Taquari River basin were compared. Variance analysis (ANOVA) was performed for the meristic data, and Principal Component Analysis (PCA) and Linear Discriminant Analysis (LDA) were conducted for morphometric data. Molecular analyses used coI, cytb, 12S and 16S mitochondrial genes, examining nucleotide diversity, haplotype diversity, genetic distance, and delimitation of possible multiple species through the Generalized Mixed Yule Coalescent (GMYC) method. Phylogenetic relationships of studied populations were also investigated through Bayesian inference. While PCA indicated a tendency of overlap between areas, ANOVA and LDA detected a subtle differentiation between populations from the two hydrographic basins. Yet, both latter analyses recovered the population from Pelotas River, a tributary to Uruguay River, as more similar to populations from Taquari River, which is congruent to morphological observations of anterior abdominal plates. The molecular data indicated a nucleotide diversity lower than the haplotypic diversity, suggestive of recent expansion. The concatenated haplotype network points to slight differentiation between areas, with each locality presenting unique and non-shared haplotypes, although with few mutational steps in general. The species delimitation by coalescence analysis suggested the presence of a variable number of OTUs depending on the inclusion or exclusion of an outgroup. In general, the morphological data suggest a subtle variation by river basin, while the genetic data indicates a weak population structuration by hydrographic areas, especially the Chapecó and Passo Fundo rivers. However, there is still not enough differentiation between the specimens to suggest multiple species. The iterative analyses indicate that Pareiorhaphis hystrix is composed of a single, although variable, species.

The Neotropical region hosts 4225 freshwater fish species, ranking first among the world's most diverse regions for freshwater fishes. Our NEOTROPICAL FRESHWATER FISHES data set is the first to produce a large‐scale Neotropical freshwater fish inventory, covering the entire Neotropical region from Mexico and the Caribbean in the north to the southern limits in Argentina, Paraguay, Chile, and Uruguay. We compiled 185,787 distribution records, with unique georeferenced coordinates, for the 4225 species, represented by occurrence and abundance data. The number of species for the most numerous orders are as follows: Characiformes (1289), Siluriformes (1384), Cichliformes (354), Cyprinodontiformes (245), and Gymnotiformes (135). The most recorded species was the characid Astyanax fasciatus (4696 records). We registered 116,802 distribution records for native species, compared to 1802 distribution records for nonnative species. The main aim of the NEOTROPICAL FRESHWATER FISHES data set was to make these occurrence and abundance data accessible for international researchers to develop ecological and macroecological studies, from local to regional scales, with focal fish species, families, or orders. We anticipate that the NEOTROPICAL FRESHWATER FISHES data set will be valuable for studies on a wide range of ecological processes, such as trophic cascades, fishery pressure, the effects of habitat loss and fragmentation, and the impacts of species invasion and climate change. There are no copyright restrictions on the data, and please cite this data paper when using the data in publications.