Microeucariotos ciliados (Alveolata, Ciliophora) encontrados em tanques de bromélias (fitotelmos): da delimitação computacional de espécies aos eventos macroevolutivos

Abstract

Phytotelmata are aquatic microhabitats formed by the accumulation of water in plant structures, such as bromeliad tanks, which harbor a wide diversity of flora, fauna, and microbiota, with ciliates being the most abundant microorganisms. Despite the increasing number of studies in these environments, knowledge about the diversity of endemic and ubiquitous species, macroevolutionary patterns, and the structure of ciliate communities is still incipient, as is the use of computational tools for species delimitation. In this context, the present study aimed to: 1 establish the macroevolutionary patterns associated with ciliate diversity in phytotelmata, estimating the timing and mode of ciliate evolution in this environment; 2 test the efficiency of computational tools in the taxonomic identification of species of the genus Tetrahymena through computational species delimitation methods: PTP and mPTP, and machine learning algorithms, to identify possible taxonomic units (ESUs) and cryptic species, as well as to infer species boundaries and cutoff points for lineage definition; and 3 molecularly characterize the community of eukaryotes associated with phytotelmata of different bromeliad species. To achieve objectives 1 and 2, maximum likelihood phylogenetic analyses were performed for the phylum Ciliophora and the genus Tetrahymena, based on the 18S and COI genes, respectively. From the inferred topology for Ciliophora, divergence times were estimated, diversification rates were calculated using the BAMM algorithm, and ancestral states were reconstructed using the MBASR package. The topology of the genus Tetrahymena was, in turn, subjected to different computational species delimitation methods. In parallel, to achieve objective 3, samples from seven bromeliad species from the Chapadão da Serra Negra Private Reserve were subjected to DNA extraction and sequencing using the PacBio platform. The generated libraries were filtered, and taxonomic characterization was performed. Additionally, various statistical analyses were carried out using the R platform. Ancestral state reconstruction analyses indicate that phytotelm-endemic ciliates colonized this environment through at least four distinct ancestral routes (freshwater, marine/brackish, terrestrial, and symbionts) and exhibit a significant increase in diversification rates. For the genus Tetrahymena, the COI gene proved highly efficient for the identification of taxonomic units, with supervised delimitation methods performing best, recovering a higher number of nominal species defined by taxonomists. Moreover, a high molecular diversity of eukaryotes was detected in phytotelmata, including previously described endemic species and significant variation among the different bromeliad species analyzed.