The objective of this study was to identify which of three types of variables, namely, mesohabitat parameters, stream margin parameters, and regional parameters, are most strongly correlated with the fish diversity of Neotropical streams located in the Upper Paraná River. Samples were collected in 28 stream stretches from the Rio das Cinzas and Paranapanema basins, Upper Paraná River, during July and August 2016, using electrofishing. Variance analysis, nonmetric multidimensional scaling (NMDS), permutational multivariate analysis of variance (PERMANOVA), and species co-occurrence (C-score) analyses were conducted to assess the influence of each environmental parameter (mesohabitats, margins, and regional factors). The co-occurrence analysis indicated the presence of non-random assemblages among mesohabitats as well, highlighting the role of biotic interactions and niche differentiation in structuring fish communities. Greater equitability was found in the riffle mesohabitat. Regarding riparian structure, silted stretches exhibited higher species abundance, with significant differences. Finally, the stream order showed significant differences in species richness, Shannon diversity, evenness, and dominance, with an evident increase in diversity values as the stream order progressed. This finding aligns with the River Continuum Concept, where increasing habitat complexity and resource availability downstream foster higher biodiversity. These results emphasize the interplay between local habitat features and broader landscape factors in shaping biodiversity patterns, highlighting the need for multi-scale approaches in conservation and management strategies for Neotropical stream ecosystems.
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