Assessment of potential reference sites for evaluating the ecological status of subtropical and temperate Brazilian lotic systems using the epilithic diatom community.

This research aimed at determining reference sites in southern Brazilian lotic systems, using the Water Quality Trophic Index (WQTI), based on epilithic diatom communities. Within the period of 2012 and 2016, 140 epilithic diatom samples from seven sampling sites within the spring area of the Andreas Stream Hydrographic Basin, Vera Cruz County, RS, Brazil, were analyzed quarterly. Similarly, from 2012 and 2014, water samples in these sampling sites were collected for determination of physicochemical and microbiological parameters, such as water temperature, pH, turbidity, dissolved oxygen, biochemical oxygen demand, nitrate, phosphate, total dissolved solids, and fecal coliforms. The results indicated the occurrence of two diatom samples groups with significant differences (p < 0.05). Group 1 (sampling sites P1, P4, and P5) was classified in the oligotrophic category with an average WQTI of 1.3 ± 0.2, while group 2 (sampling sites P2, P3, P6, and P7) was classified in the ß-mesotrophic category with an average value of 2.0 ± 0.4). Achnanthidium minutissimum and Platessa hustedtii the dominant species were classified as oligotrophic taxa. Thus, we proposed the upper limit of the first interval of the WQTI score scale, equal to 1.5, as a reference value to differentiate the limits between the "high" and "good" ecological status to determine "reference sites" for subtropical and temperate Brazilian aquatic systems. The physical, chemical, and microbiological water quality evaluation gives support to this proposal, as far as the results indicated a significant improvement in the water quality, classifying sampling sites P1, P4, and P5 as having good quality appropriate for multiple uses.

Genetic barcoding of Ecuadorian epilithic diatom species suitable as water quality bioindicators.

Diatom identification is a key step in using these microorganisms as water quality bioindicators. Morphological diagnosis is a difficult task due to the enormous number of species and their microscopic size. This can be overcome using molecular tools to complement the diagnosis. The main goal of this work was to obtain the DNA barcode of Ecuadorian epilithic diatoms with a wide geographical distribution, a well-defined ecological range and characteristics that allow them to be reliable indicator species. Unialgal diatom cultures were obtained from environmental samples of Ecuadorian Andean streams. Morphological characterization of cultures was carried out under SEM microscopy. For molecular characterization, 18SV4 and rbcL barcodes were sequenced from each strain and blasted against a GenBank database. A phylogenetic tree for each barcode was constructed using the ML method including sequences of strains of the studied species from different geographical locations. The results showed the following five species to be suitable as bioindicators and these were isolated. Sellaphora seminulum (strain JA01b, c), Nitzschia fonticola (strain SP02a) and N. palea (strain CA01a) are tolerant to eutrophication; Eolimna minima (strain CH02a) is a mesotrophic water bioindicator, and Achnanthidium minutissimum (strain JA01a) is an oligotrophic water bioindicator. The comparison with the GenBank database of the barcoding regions supported the morphological identification. The barcoding sequences of the strains showed a high percentage of identity with the sequences reported in INSDC databases for the same species. The topology of the phylogenetic trees demonstrates that epilithic diatoms from Ecuador are closely related to those of same species isolated from other geographical regions. This study is a first attempt to establish a morphological and molecular taxonomic reference library for neotropical diatoms. This study demonstrates that it would be feasible to use the existing barcoding data for diatoms to develop molecular tools for the bioassessment of aquatic ecosystems in the Ecuadorian Andean region.

L'identification des diatomées est une étape clé dans l'utilisation de ces microorganismes comme bioindicateurs de la qualité de l'eau. Le diagnostic morphologique est une tâche difficile en raison du nombre considérable d'espèces et de leur dimension microscopique. Il est possible de surmonter cette difficulté en utilisant des techniques moléculaires pour compléter le diagnostic. L'objectif principal de ce travail était d'obtenir le code-barre de l'ADN des diatomées épilithiques équatoriennes ayant une large distribution géographique, une niche écologique bien définie et des caractéristiques leur permettant d'être des espèces indicatrices fiables. Des cultures de diatomées unialgales ont été obtenues à partir d'échantillons environnementaux de cours d'eau des Andes équatoriennes. La caractérisation morphologique des cultures a été réalisée sous microscopie MEB. Pour la caractérisation moléculaire, les codes-barres 18SV4 et rbcL ont été séquencés à partir de chaque souche et comparés à la base de données GenBank. Pour chaque code-barres, un arbre phylogénétique a été construit à partir de la méthode ML comprenant des séquences de souches des espèces étudiées, provenant de différents lieux géographiques. Les résultats ayant montré que les cinq espèces suivantes étaient appropriées comme bioindicateurs, elles ont été isolées. Sellaphora seminulum (souche JA01b, c), Nitzschia fonticola (souche SP02a) et N. palea (souche CA01a) sont tolérantes à l'eutrophisation ; Eolimna minima (souche CH02a) est un bioindicateur d'eau mésotrophe, et Achnanthidium minutissimum (souche JA01a) est un bioindicateur d'eau oligotrophe. La comparaison avec la base de données GenBank des régions de code-barres a supporté leurs identifications morphologiques. Les séquences de code-barres des souches ont montré un pourcentage élevé d'identité génétique avec les séquences signalées dans les bases de données de l'INSDC pour la même espèce. La topologie des arbres phylogénétiques démontre que les diatomées épilithiques de l'Équateur sont étroitement liées à celles des mêmes espèces isolées d'autres régions géographiques. Cette étude est une première tentative d'établir une bibliothèque de référence morphologique et taxonomique moléculaire pour les diatomées néotropicales. Cette étude démontre qu'il serait possible d'utiliser les données de code-barres existantes pour les diatomées afin de développer des instruments moléculaires pour la bioévaluation des écosystèmes aquatiques dans la région andine équatorienne.

Response of epilithic diatom communities to environmental gradients along an Ecuadorian Andean River.

Epilithic diatom communities are particularly suitable for the evaluation of freshwater quality. In Ecuador, however, no water quality index includes this biotic parameter. This work is the first attempt in the country to determine the composition of epilithic diatom communities associated with different degrees of eutrophication. This was accomplished by measuring physical, chemical and microbiological variables at five sampling sites along the Pita River, Ecuador, from August to December 2016. The results indicate a clear gradient of eutrophication from sampling sites P1 (good water) in the upper reaches to P5 (bad water) located in the lower reaches. Concerning diatom analyses, the results indicated a high diversity for tropical areas in terms of species richness, varying from S=34 in headwaters to S=42 downstream. Moreover, the results obtained suggest a lack of concordance with the trophic values given to some of the epilithic diatoms in the literature. There were also species that seem to be sensitive to downstream nutrient increases that were not considered as bioindicators in previous studies. We concluded that the trophic values of diatom species available in the scientific literature are not directly applicable to their sites in the Pita River. Hence, it is necessary to establish a trophic diatom index for the Andean region of Ecuador.

Development of the Trophic Water Quality Index (TWQI) for subtropical temperate Brazilian lotic systems.

This research aimed at developing the Trophic Water Quality Index (TWQI) for subtropical temperate Brazilian lotic systems based on a review of the indicative values of diatom species obtained using multivariate analysis techniques and considering the environmental gradient defined by a series of measured physical, chemical, and microbiological variables. Sampling was conducted from 2005 to 2009 in the Pardo River Hydrographic Basin, Rio Grande do Sul (RS), Brazil, and in the Andreas Stream Hydrographic Basin, RS, from 2012 to 2013. A total of 140 biological samples and 211 abiotic samples were collected. Data were analyzed by cluster analysis based on the Ward method and canonical correspondence analysis (CCA). The results indicated that total phosphate, turbidity, ammonia nitrogen, electrical conductivity, dissolved oxygen, and thermotolerant coliforms showed a significant correlation with the sample ordination made by CCA, in relation to a gradient of eutrophication. Eutrophication was operationally defined in a broad sense, including the problem of organic pollution and eutrophication of the water. The determination of the different tolerance degrees to eutrophication of the diatom taxa was used to assign trophic values of 1, 2.5, and 4 to species, corresponding to levels of low, medium, and high tolerance, respectively. By using the trophic values obtained for each diatom species, the TWQI constituted a new technological tool for environmental monitoring studies and showed a consistent, robust, and objective database for water quality assessment in subtropical temperate Brazilian lotic systems.