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Amazonia harbours a vast biotic and ecological diversity, enabling investigation of the effects of microevolutionary processes and environmental variation on species diversification. Integrative approaches combining phenotypic and genetic variation can improve our knowledge on diversification processes in megadiverse regions. Here, we investigate the influence of environmental and geographic variation on the genetic and morphological differentiation in the Amazonian Boana calcarata-fasciata (Anura: Hylidae) species complex. We analysed the variation of one mtDNA gene from individuals of different forest environments, and assessed their phylogenetic relationships and species limits to define the lineages to perform a phenotypic-environmental approach. We collected morphological data (head shape and size) using 3D models and investigated the phylogenetic signal, evolutionary model and influence of environmental variables on morphology. We verified associations between environmental and geographical distances with morphological and genetic variation using distance-based redundancy analyses and Mantel tests. We found an even higher cryptic diversity than already recognized within the species complex. Body size and head shape varied among specimens, but did not present phylogenetic signal, diverging under a selective evolutionary model. Our results show that diverse factors have influenced morphological and genetic variation, but environmental conditions such as vegetation cover, precipitation and climate change velocity influenced morphological diversification. Possible population-level mechanisms such as parallel morphological evolution or plastic responses to similar environments could account for such patterns in these typical Amazonian treefrogs.

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Sampling gaps across the logistically challenging and extremely biodiverse Amazonia largely hamper our understanding of broad-scale amphibian and reptile diversity patterns in this ecosystem. The Juruá River basin, a southwestern tributary of the Amazon River, is one of these undersampled areas, with only punctual information documented for these vertebrates that are spatially or temporally biased. This is especially the case for the lower-middle courses of the Juruá River, which also has comparatively less protected areas than its upper course. In order to fill some biodiversity knowledge gaps associated with amphibians and reptiles in this river basin, we combined results of our field expeditions carried out in 1992, 2005-2007, and 2018 to the Reserva Extrativista (Extractive Reserve) do Baixo Juruá, a Brazilian protected area in the right bank of the lower Juruá River. Amphibians and reptiles were sampled using four complementary methods: active surveys, pitfall traps, funnel traps, and trammel nets. We identified species or updated their taxonomic status with a reanalysis of the external morphology of the preserved material in the light of novel taxonomic literature (more than 1,500 specimens) and employment of DNA barcoding analyses for some newly collected specimens with contentious taxonomic status. Our combined sampling evidenced 149 species of amphibians and reptiles occurring in this protected area (72 amphibians, 68 squamates, six chelonians, and three crocodilians). Recorded species highlight the value of the lower Juruá River region as harboring quite diverse assemblages for these vertebrates, with species typical of the western and southwestern Amazonia sub-regions. Remarkable species records are presented, as well as accounts on species with lower taxonomic resolution. Furthermore, we discuss the biogeographic affinities of recorded assemblages based on the species geographic range and preferred habitats, and the value of this protected area to preserve the regional biological diversity.

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